Microbial Metabolism

Microbial Metabolism How do pathogens acquire energy and nutrients at the expense of a patient’s health? How does grape juice turn into wine, and how does yeast cause bread to rise? When laboratory personnel perform biochemical tests to identify unknown microorganisms and help diagnose disease, what exactly are they doing? The answers to all of these questions require an understanding of microbial metabolism,1 the collection of controlled biochemical reactions that takes place within a microbe. Although it is true that metabolism in its entirety is complex, consisting of thousands of chemical reactions and control mechanisms, the reactions are nevertheless elegantly logical and can be understood in a simplified form. In this chapter, we concern ourselves only with central metabolic pathways and energy metabolism. Your study of metabolism will be manageable if you keep in mind that the ultimate function of an organism’s metabolism is to reproduce the organism and that metabolic processes are guided by the following eight elementary statements: • Every cell acquires nutrients, which are the chemicals necessary as building blocks and energy sources for metabolism. • Metabolism requires energy from light or from the catabo­ lism (kă-tab´ō-lizm; breakdown) of acquired nutrients. • Energy is often stored in the chemical bonds of adenosine triphosphate (ATP). • Using enzymes, cells catabolize nutrient molecules to form elementary building blocks called precursor metabolites. • Using precursor metabolites, other enzymes, and energy from ATP, cells construct larger building blocks in anabolic (an-ă-bol´ik; biosynthetic) reactions. • Cells use enzymes and additional energy from ATP to anabolically link building blocks together to form macromolecules in polymerization reactions. • Cells grow by assembling macromolecules into cellular structures such as ribosomes, membranes, and cell walls. • Cells typically divide into two once they have doubled in size. We will discuss each aspect of metabolism in the chapters that most directly apply. (For instance, Chapter 3 discussed the first step of metabolism—the active and passive transport of nutrients into cells.) The current chapter examines the importance of enzymes in catabolic and anabolic reactions, studies three ways that ATP molecules are synthesized, and shows that catabolic and anabolic reactions are linked. We also examine the catabolism of nutrient molecules; the anabolic reactions involved in the synthesis of carbohydrates, lipids, amino acids, and nucleotides; and a few ways that cells control their metabolic activities. (Chapter 7 discusses genetic control of metabolism and the polymerization of DNA, RNA, and proteins, and Chapters 11 and 12 cover specifics of cell division.) 1From Greek metabole, meaning “change.” M05_BAUM7206_05_SE_C05.indd 125       Basic Chemical Reactions Underlying Metabolism Basic Chemical Reactions Underlying Metabolism In the following sections, we will examine the basic concepts of catabolism, anabolism, and a special class of reactions called oxidation­reduction reactions. The latter involve the transfer of electrons and energy carried by electrons between molecules. Then we will turn our attention briefly to the synthesis of ATP and energy storage before we discuss the organic catalysts called enzymes, which make metabolism possible. ▶ ANIMATIONS: Metabolism: Overview Catabolism and Anabolism LE A R N I N G O UTC O M E 5.1 Distinguish among metabolism, anabolism, and catabolism. Metabolism, which is all of the chemical reactions in an organism, can be divided into two major classes of reactions: catabolism and anabolism (FIGURE 5.1). A series of such reactions is called a pathway. Cells have catabolic pathways, which break larger molecules into smaller products, and anabolic pathways, which synthesize large molecules from smaller ones. Even though catabolic and anabolic pathways are intimately linked in cells, it is often useful to study the two types of pathways as though they were separate. When catabolic pathways break down large molecules, they release energy; that is, catabolic pathways are exergonic (ek-sergon´ik). Cells store some of this released energy in the bonds of ATP, though much of the energy is lost as heat. Another result Energy lost as heat Energy lost as heat ATP Energy stored ISM OL B TA CA Nutrients Energy used ANABOLISM Precursor molecules ADP Energy storage (carbohydrates, lipids, etc.) Larger building blocks Macromolecules Cellular structures Cellular (membranes, processes ribosomes, etc.) (cell growth, cell division, etc.) ▲ FIGURE 5.1 Metabolism is composed of catabolic and anabolic reactions. Catabolic reactions are exergonic—they release energy, some of which is stored in ATP molecules, though most of the energy is lost as heat. Anabolic reactions are endergonic—they require energy, typically provided by ATP. There is some heat loss in anabolism as well. The products of catabolism provide many of the building blocks (precursor metabolites) for anabolic reactions. These reactions produce macromolecules and cellular structures, leading to cell growth and division. 11/23/16 3:00 AM r hapt       e 5   Microbial Metabolism of the breakdown of large molecules by catabolic pathways is the production of numerous smaller molecules, some of which are precursor metabolites of anabolism. Some organisms, such as Escherichia coli (esh-ĕ-rik´ē-ăkō´īlī), can synthesize everything in their cells from precursor metabolites alone; other organisms must acquire at least some of their anabolic building blocks from outside themselves as nutrients. Catabolic pathways, but not necessarily individual catabolic reactions, produce ATP, or metabolites, or both. An example of a catabolic pathway is the breakdown of lipids into glycerol and fatty acids. Anabolic pathways are functionally the opposite of catabolic pathways in that anabolic pathways synthesize macromolecules and cellular structures. Because building anything requires energy, anabolic pathways are endergonic (en- dergon´ik); that is, they require more energy than they release. The energy required for anabolic pathways usually comes from ATP molecules produced during catabolism. An example of an anabolic pathway is the synthesis of cell membrane lipids from glycerol and fatty acids. To summarize, a cell’s metabolism involves both catabolic pathways that break down macromolecules to supply molecular building blocks and energy in the form of ATP, and anabolic pathways that use the building blocks and ATP to synthesize macromolecules needed for growth and reproduction. Oxidation and Reduction Reactions LE A R N I N G O U TC O M E 5.2 Contrast oxidation and reduction reactions. ­ ­ Many metabolic reactions involve the transfer of electrons, and keeping track of these electrons is important for your understanding of metabolism. Why is it important? Electrons carry energy, and you need to understand how a cell uses that energy. Electrons are transferred from an electron donor (a molecule that donates an electron) to an electron acceptor (a molecule that accepts an electron). Such electron transfers are called oxidation-reduction reactions, or redox reactions (FIGURE 5.2) . An electron acceptor is said to be reduced. This may seem backward, but electron acceptors are reduced because their gain in electrons reduces their overall electrical charge (i.e., they are more negatively charged). Molecules that lose electrons are said to be oxidized because in many reactions their electrons are donated to oxygen atoms. An acronym to help you remember these concepts is OIL RIG: oxidation involves loss; reduction involves gain. Reduction and oxidation reactions always happen simultaneously because every electron donated by one chemical is accepted by another chemical. A chemical may be reduced by gaining either a simple electron or an electron that is part of a hydrogen atom—which is composed of one proton and one electron. In contrast, a molecule may be oxidized in one of three ways: by losing a simple electron, by losing a hydrogen atom, or by gaining an oxygen atom. Biological oxidations often involve the loss of hydrogen atoms; such reactions are also called dehy­ drogenation (dē-hī´drō-jen-ā´shŭn) reactions. M05_BAUM7206_05_SE_C05.indd 126 Reduction Oxidized donor Electron donor e– e– Electron acceptor Reduced acceptor Oxidation ▲ FIGURE 5.2 Oxidation-reduction, or redox, reactions. When electrons are transferred from donor molecules to acceptor molecules, donors become oxidized, and acceptors become reduced. Why are acceptor molecules said to be reduced when they are gaining electrons? Figure 5.2 Reduction refers to the overall electrical charge on a molecule. Because electrons have a negative charge, the gain of an electron reduces the molecule’s overall charge. C Electrons rarely exist freely in cytoplasm; instead, they orbit atomic nuclei. Therefore, cells use electron carrier molecules to carry electrons (often in hydrogen atoms) from one location in a cell to another. Three important electron carrier molecules are nicotinamide adenine dinucleotide (NAD + ), nicotinamide adenine dinucleotide phosphate (NADP + ), and flavin adenine dinucleotide (FAD). These molecules are derived from vitamins. Cells use each of these electron carriers in specific metabolic pathways to carry pairs of electrons. One of the electrons carried by either NAD + or NADP + is part of a hydrogen atom, forming NADH or NADPH. FAD carries two electrons as hydrogen atoms (FADH2). Many metabolic pathways, including those that synthesize ATP, require electron carrier molecules. ▶ ANIMATIONS: Oxidation-Reduction Reactions ATP Production and Energy Storage LE A R N I N G O UTC O M E 5.3 Compare and contrast the three types of ATP phosphorylation. Nutrients contain energy, but that energy is spread throughout their chemical bonds and generally is not concentrated enough for use in anabolic reactions. During catabolism, organisms release energy from nutrients that can then be concentrated and stored in high-energy phosphate bonds of molecules such as ATP. This happens by a general process called phosphoryla­ tion (fos´fōr-i-lā´shŭn), in which inorganic phosphate ((PO4 3- ) is added to a substrate. For example, cells phosphorylate adenosine diphosphate (ADP), which has two phosphate groups, to form adenosine triphosphate (ATP), which has three phosphate groups (see Chapter 2, Figure 2.27). Why is ATP well suited to serve as the primary short-term energy carrier in metabolic pathways? First, it is multifunctional as a ribonucleotide for use in synthesizing RNA. Second, it is highly water soluble and can accumulate to high concentrations in cells with no ill effects. Third, it has two different levels of energy 11/23/16 3:00 AM       Basic Chemical Reactions Underlying Metabolism • Substrate-level phosphorylation (see pp. 134–135), which involves the transfer of phosphate to ADP from another phosphorylated organic compound • Oxidative phosphorylation (see pp. 139–140), in which energy from redox reactions of respiration (described shortly) is used to attach inorganic phosphate to ADP • Photophosphorylation (see pp. 145–146), in which light energy is used to phosphorylate ADP with inorganic phosphate We will investigate each of these in more detail as we proceed through the chapter. After ADP is phosphorylated to produce ATP, anabolic pathways use some energy of ATP by breaking a phosphate bond (which re-forms ADP). Thus, the cyclical interconversion of ADP and ATP functions somewhat like rechargeable batteries: ATP molecules store energy from catabolic reactions (and from light in photosynthetic organisms) and then release stored energy to drive cellular processes (including anabolic reactions, active transport, and movement). ADP molecules can be “recharged” to ATP again and again. Cells use ATP as a very short-term energy storage molecule; the average half-life of a molecule of ATP is measured in seconds. The Roles of Enzymes in Metabolism LE A R N I N G O U TC O M E S 5.4 Make a table listing the six basic types of enzymes, their activities, and an example of each. 5.5 Describe the components of a holoenzyme, and contrast protein and RNA enzymes. 5.6 Define activation energy, enzyme, apoenzyme, cofactor, coen­ zyme, active site, and substrate, and describe their roles in enzyme activity. 5.7 Describe how temperature, pH, substrate concentration, and competitive and noncompetitive inhibition affect enzyme activity. Chemical reactions occur when bonds are broken or formed between atoms. In catabolic reactions, a bond must be destabilized before it will break, whereas in anabolic reactions, reactants collide with sufficient energy for bonds to form between them. In anabolism, increasing either the concentrations of reactants or ambient temperatures increases the number of collisions and produces more chemical reactions; however, in living organisms, neither reactant concentration nor temperature is usually high enough to ensure that bonds will form. Therefore, the chemical reactions of life depend on catalysts, which are chemicals that increase the likelihood of a reaction but are not permanently changed in the process. Organic catalysts are known as enzymes. ▶ ANIMATIONS: Enzymes: Overview M05_BAUM7206_05_SE_C05.indd 127 Naming and Classifying Enzymes The names of enzymes usually end with the suffix -ase, and the name of each enzyme often incorporates the name of that enzyme’s substrate, which is the molecule the enzyme acts on. Based on their mode of action, enzymes can be grouped into six basic categories: • Hydrolases catabolize molecules by adding water in a decomposition process known as hydrolysis. Hydrolases are used primarily in depolymerizing catabolism of macromolecules. • Isomerases2 rearrange atoms within a molecule but do not add or remove anything (so they are neither catabolic nor anabolic). • Ligases, or polymerases, join two molecules together (and are thus anabolic). They often use energy supplied by ATP. • Lyases split large molecules (and are thus catabolic) without using water in the process. • Oxidoreductases remove electrons from (oxidize) or add electrons to (reduce) various substrates. They are used in both catabolic and anabolic pathways. • Transferases transfer functional groups, such as an amino group (NH2), a phosphate group (PO4 3- ), or a two-carbon (acetyl) group, between molecules. Transferases can be anabolic. TABLE 5.1 on p. 128 summarizes these types of enzymes and gives examples of each. The Makeup of Enzymes Many protein enzymes are complete in themselves, but others are composed of both protein and nonprotein portions. The proteins in these combinations are called apoenzymes (ap´ōen-zīms). Apoenzymes are inactive if they are not bound to one or more of the nonprotein substances called cofactors. Cofactors are either inorganic ions (such as iron, magnesium, zinc, or copper ions) or certain organic molecules called coenzymes (ko-en´zīms). All coenzymes are either vitamins or contain vitamins, which are organic molecules that are required for metabolism but cannot be synthesized by certain organisms (especially mammals). Some apoenzymes bind with inorganic cofactors, some bind with coenzymes, and some bind with both. The binding of an apoenzyme and its cofactor(s) forms an active enzyme, called a holoenzyme (hol-ō-en´zīm; FIGURE 5.3). TABLE 5.2 on p. 128 lists several examples of inorganic cofactors and organic cofactors (coenzymes). Note that three important coenzymes are the electron carriers NAD + , NADP + , and FAD, which, as we have seen, carry electrons as part of hydrogen atoms from place to place within cells. We will examine more closely the roles of these coenzymes in the generation of ATP later in the chapter. ­ donation, ATP to ADP and ATP to AMP, depending on what is needed for a reaction. Fourth, ATP also can also serve as a phosphate donor. As we will examine in the following sections, cells phosphorylate ADP to form ATP in three specific ways: 2An isomer is a compound with the same molecular formula as another molecule, but with a different arrangement of atoms. 11/23/16 3:00 AM r e 5   Microbial Metabolism hapt       C TABLE 5.1 Enzyme Classification Based on Reaction Types Class Type of Reaction Catalyzed Example Hydrolase Hydrolysis (catabolic) Lipase—breaks down lipid molecules Isomerase Rearrangement of atoms within a molecule (neither catabolic nor anabolic) Phosphoglucoisomerase—converts glucose 6-phosphate into fructose 6-phosphate during glycolysis Ligase or Polymerase Joining two or more chemicals together (anabolic) Acetyl-CoA synthetase—combines acetate and coenzyme A to form acetyl-CoA for the Krebs cycle Lyase Splitting a chemical into smaller parts without using water (catabolic) Fructose-1,6-bisphosphate aldolase—splits fructose 1,6-bisphosphate into G3P and DHAP Oxidoreductase Transfer of electrons or hydrogen atoms from one molecule to another Lactic acid dehydrogenase—oxidizes lactic acid to form pyruvic acid during fermentation Transferase Moving a functional group from one molecule to another (may be anabolic) Hexokinase—transfers phosphate from ATP to glucose in the first step of glycolysis Inorganic cofactor Active site Coenzyme (organic cofactor) Apoenzyme (protein) Holoenzyme ▲ FIGURE 5.3 Makeup of a protein holoenzyme. The com- bination of a proteinaceous apoenzyme with one or more cofactors forms a holoenzyme, which is the active form. A cofactor is either an inorganic ion or a coenzyme, which is an organic cofactor derived from a vitamin. An apoenzyme is inactive unless it is bound to its cofactors. Name four metal ions that can act as cofactors. Figure 5.3 Iron, magnesium, zinc, and copper ions can act as cofactors. TABLE 5.2 Not all enzymes are proteins; some are RNA enzymes called ribozymes. In eukaryotes, ribozymes process other RNA molecules by removing sections of RNA and splicing the remaining pieces together. Recently, researchers have discovered that a ribozyme makes up the functional core of a ribosome, which is a cell’s protein factory; in other words, all protein enzymes are made by ribozymes. Enzyme Activity Within cells, enzymes catalyze reactions by lowering the activation energy, which is the amount of energy needed to trigger a chemical reaction (FIGURE 5.4). Heat can provide energy to trigger reactions, but for most metabolic reactions the temperatures needed to reach activation energy are often too high to allow cells to survive. So, for metabolism to occur, enzymes are needed. This is true regardless of whether the enzyme is a protein or RNA or whether the chemical reaction is anabolic or catabolic. Representative Cofactors of Enzymes Examples of Use in Enzymatic Activity Substance Transferred in Enzymatic Activity Vitamin Source (of Coenzyme) Forms bond with ADP during phosphorylation Phosphate None Nicotinamide adenine dinucleotide (NAD + ) Carrier of reducing power Two electrons and a hydrogen ion Niacin (B 3) Nicotinamide adenine dinucleotide phosphate (NADP + ) Carrier of reducing power Two electrons and a hydrogen ion Niacin (B 3) Flavin adenine dinucleotide (FAD) Carrier of reducing power Two hydrogen atoms Riboflavin (B 2) Tetrahydrofolate Used in synthesis of nucleotides and some amino acids One-carbon molecule Folic acid (B 9) Coenzyme A Formation of acetyl-CoA in Krebs cycle and beta-oxidation Two-carbon molecule Pantothenic acid (B 5) Pyridoxal phosphate Transaminations in the synthesis of amino acids Amine group Pyridoxine (B 6) Thiamine pyrophosphate Decarboxylation of pyruvic acid Aldehyde group (CHO) Thiamine (B 1) Cofactors Inorganic (Metal Ion) Magnesium (Mg2+ ) Organic (Coenzymes) M05_BAUM7206_05_SE_C05.indd 128 11/23/16 3:00 AM       Basic Chemical Reactions Underlying Metabolism E nzymes catalyze reactions by lowering the activation energy—that is, the energy needed to trigger the reaction. Does this graph represent a catabolic or an anabolic reaction? Figure 5.4 Energy of the products is lower than energy of the reactants. Thus, the graph represents catabolism. The activity of enzymes depends on the closeness of fit between the functional sites of an enzyme and its substrate. The shape of an enzyme’s functional site, called its active site, is complementary to the shape of the enzyme’s substrate. Generally, the shapes and locations of only a few amino acids in a protein enzyme or nucleotides in a ribozyme determine the shape of that enzyme’s active site. A change in a single component—for instance, through mutation—can render the enzyme less effective or even completely nonfunctional. Enzyme­substrate specificity, which is critical to enzyme activity, has been likened to the fit between a lock and key. This analogy is not completely apt because enzymes’ active sites change shape slightly when they bind to their substrate, almost as if a lock could grasp its key once it had been inserted. This latter description of enhanced enzyme-substrate specificity is called the induced-fit model (FIGURE 5.5). 1 An enzyme associates with its specific substrate molecule, which has a shape complementary to that enzyme’s active site. 2 The enzyme and its substrate bind to form a temporary intermediate compound called an enzyme­substrate com­ plex. The binding of the substrate induces the enzyme to fit the shape of the substrate even more closely—the induced-fit model. 3 Bonds within the substrate are broken, forming two or more products in catabolic reactions. (In anabolic reactions, reactants are linked together to form products.) 4 The enzyme dissociates from the newly formed products, which diffuse away from the site of the reaction. 5 The enzyme resumes its original configuration and is ready to associate with another substrate molecule.  ▲ FIGURE 5.4 The effect of enzymes on chemical reactions.  Progress of reaction  Products  Energy Reactants  Activation energy without enzyme Activation energy with enzyme In some cases, several different enzymes possess active sites that are complementary to various portions of a single substrate molecule. For example, an important precursor metabolite called phosphoenolpyruvic acid (PEP) is the substrate for at least five enzymes. Depending on the enzyme involved, various products are produced from PEP. In one catabolic pathway, PEP is converted to pyruvic acid, whereas in a particular anabolic pathway, PEP is converted to the amino acid phenylalanine. Although the exact ways that enzymes lower activation energy are not known, it appears that several mechanisms are involved. Some enzymes appear to bring reactants into sufficiently close proximity to enable a bond to form, whereas other enzymes change the shape of a reactant, inducing a bond to be broken. In any case, enzymes increase the likelihood that bonds will form or break. The activity of enzymes is believed to follow the process illustrated in FIGURE 5.6, which depicts the catabolic lysis of a molecule called fructose 1,6-bisphosphate: Many factors influence the rate of enzymatic reactions, including temperature, pH, enzyme and substrate concentrations, and the presence of inhibitors. ▶ ANIMATIONS: Enzymes: Steps in a Reaction Substrate Active sites similar to substrate’s shape Enzyme Substrate binding induces an enzyme’s active site to more closely match the shape of the substrate (induced fit) ▲ FIGURE 5.5 Enzymes fitted to substrates. The induced-fit model of enzyme-substrate interaction. An enzyme’s active site is generally complementary to the shape of its substrate, but a perfect fit between them does not occur until the substrate and enzyme bind to form a complex, at which time the enzyme’s active site changes to fit the substrate more closely. M05_BAUM7206_05_SE_C05.indd 129 Temperature As mentioned, higher temperatures tend to increase the rate of most chemical reactions because molecules are moving faster and collide more frequently, encouraging bonds to form or break. However, this is not entirely true of enzymatic reactions because as the temperature rises above the stability point for the enzyme, the enzyme loses structure and the active site is lost. Each enzyme has an optimal temperature for its activity (FIGURE 5.7a). The optimum temperature for the enzymes in the human body is 37°C, which is normal body temperature. Part of the reason certain pathogens can cause disease in humans is that the optimal temperature for the enzymes in those microorganisms is also 37°C. The enzymes of some other microorganisms, however, function best at different temperatures; this is the case for hyperthermophiles, organisms that grow best at temperatures above 80°C. 11/23/16 3:00 AM r e 5   Microbial Metabolism hapt       C Substrate (Fructose 1,6-bisphosphate) O CH2 H O H O P OH OH Enzyme (Fructose-1,6bisphosphate aldolase) CH2 OH Enzyme activity P H 1 O P H2C H 2 5 H2C H H OH OH OH H O 10 15 20 25 30 35 40 45 50 Temperature (5C) P (a) Temperature O CH2 H OH OH OH H O P Enzyme activity O CH2 Enzymesubstrate complex 3 P O 4 Glyceraldehyde-3P P O CH2 Dihydroxyacetone-P CH2 HCOH CHO C O 0 P 2 4 6 8 10 pH (b) pH O CH2OH Products Enzyme activity ▲ FIGURE 5.6 The process of enzymatic activity. Shown here is the lysis of fructose 1,6-bisphosphate by the enzyme fructose-1,6-bisphosphate aldolase (a catabolic reaction). After the enzyme associates with the substrate 1 , the two molecules bind to form an enzyme-substrate complex. 2 As a result of binding, the enzyme’s active site is induced to fit the substrate even more closely; then bonds within the substrate are broken 3 , after which the enzyme dissociates from the two products 4 . The enzyme resumes its initial shape and is then ready to associate with another substrate molecule 5 . This entire process occurs 14 times per second at 37°C (body temperature) for a single enzyme. Saturation point Substrate concentration (c) Substrate concentration M05_BAUM7206_05_SE_C05.indd 130 E ffects on each enzyme will vary. (a) Rising temperature enhances enzymatic activity to a point, but above some optimal temperature an enzyme denatures and loses function. (b) nzymes typically have some optimal pH, at which point enzymatic activity reaches a maximum. (c) At lower substrate concentrations, enzyme activity increases as the substrate concentration increases and as more and more active sites are utilized. At the substrate concentration at which all active sites are utilized, termed the saturation point, enzymatic activity reaches a maximum, and any additional increase in substrate concentration has no effect on enzyme activity. What is the optimal pH of the enzyme shown in part (b)? E ­ pH Extremes of pH also denature enzymes when ions released from acids and bases interfere with hydrogen bonding and ▲ FIGURE 5.7 Representative effects of temperature, pH, and substrate concentration on enzyme activity. Figure 5.7 The enzyme’s optimal pH is approximately 7.2. If temperature rises beyond a certain critical point, the noncovalent bonds within an enzyme (such as the hydrogen bonds between amino acids) will break, and the enzyme will denature (FIGURE 5.8). Denatured enzymes lose the specific three- dimensional structure of the active site, so they are no longer functional. Denaturation is said to be permanent when an enzyme cannot regain its original structure once conditions return to normal, much like the irreversible solidification of the protein albumin when egg whites are cooked and then cooled. In other cases, denaturation is reversible—the denatured enzyme’s noncovalent bonds re-form on the return of normal conditions. 11/23/16 3:00 AM       Basic Chemical Reactions Underlying Metabolism Distorted, nonfunctional active site Substrate Active site becomes functional Allosteric site Allosteric activator ▲ FIGURE 5.9 Allosteric activation. Allosteric activation results when the binding of an activator molecule to an allosteric site causes a change in the active site that makes it capable of binding substrate. Functional protein Denatured protein ▲ FIGURE 5.8 Denaturation of protein enzymes. Breakage of noncovalent bonds (such as hydrogen bonds) causes the protein to lose its secondary and tertiary structure and become denatured; as a result, the enzyme is no longer functional. distort and disrupt an enzyme’s secondary and tertiary structures. Therefore, each enzyme has an optimal pH (FIGURE 5.7b). Changing the pH provides a way to control the growth of unwanted microorganisms by denaturing their proteins. For example, vinegar (acetic acid, pH 3.0) acts as a preservative in dill pickles, and ammonia (pH 11.5) can be used as a disinfectant. Enzyme and Substrate Concentration Another factor that determines the rate of enzymatic activity within cells is the concentration of substrate present (FIGURE 5.7c). As substrate concentration increases, enzymatic activity increases as more and more enzyme active sites bind more and more substrate molecules. Eventually, when all enzyme active sites have bound substrate, the enzymes have reached their saturation point, and the addition of more substrate will not increase the rate of enzymatic activity. Obviously, the rate of enzymatic activity is also affected by the concentration of enzyme within cells. In fact, one way that organisms regulate their metabolism is by controlling the quantity and timing of enzyme synthesis. In other words, many enzymes are produced in the amounts and at the times they are needed to maintain metabolic activity. (Chapter 7 discusses the role of genetic mechanisms in regulating enzyme synthesis.) Additionally, eukaryotic cells control some enzymatic activities by compartmentalizing enzymes inside membranes so that certain metabolic reactions proceed physically separated from the rest of the cell. For example, white blood cells catabolize phagocytized pathogens using enzymes packaged within lysosomes. Control of Enzymatic Activity Organisms can influence enzymatic activity to control metabolic activity. Here, we consider both positive and negative control. An enzyme can be activated by the binding of a cofactor to the enzyme at a site located away from the active site—a site called an allosteric (al-ō-stār´ik) site. In allosteric activation, the binding of an activator, such as a heavy-metal ion or other cofactor, to an allosteric site causes the enzyme’s active site to change shape, which activates the enzyme (FIGURE 5.9). M05_BAUM7206_05_SE_C05.indd 131 Substances that block enzyme activity are called inhibitors. Enzymatic inhibitors may be either competitive or noncompetitive. Competitive inhibitors are shaped such that they fit into an enzyme’s active site and thus prevent the normal substrate from binding (FIGURE 5.10a). Such inhibitors do not undergo a chemical reaction to form products. Competitive inhibitors can bind permanently or reversibly to an active site. Permanent binding results in permanent loss of enzymatic activity; reversible competition can be overcome by an increase in the concentration of substrate molecules, increasing the likelihood that active sites will be filled with substrate instead of inhibitor (FIGURE 5.10b). ▶ ANIMATIONS: Enzymes: Competitive Inhibition An example of competitive inhibition is the action of the antibacterial drug sulfanilamide, which has a shape similar to that Substrate Competitive inhibitor Enzyme (a) Substrate Reversible competitive inhibitor Enzyme Increase in substrate concentration (b) ▲ FIGURE 5.10 Competitive inhibition of enzyme activity. (a) Inhibitory molecules, which are similar in shape to substrate molecules, compete for and block active sites. (b) Reversible inhibition can be overcome by an increase in substrate concentration. 11/23/16 3:00 AM r e 5   Microbial Metabolism hapt       C of a metabolite called para-aminobenSO2NH2 COOH zoic acid (PABA). Sulfanilamide can fill the active site of an enzyme required for the conversion of PABA into folic acid. Folic acid is essential for RNA and DNA synthesis. Once sulfanilamide is NH2 NH2 bound to the enzyme, it stays bound PABA and inhibits synthesis of folic acid and Sulfanilamide thus prevents synthesis of RNA and DNA. Thus, sulfanilamide effectively inhibits microbes that make their own folic acid for synthesizing nucleic acids. Humans do not synthesize folic acid—we must acquire it as a vitamin in our diets—so sulfanilamide does not affect us in this way. Noncompetitive inhibitors do not attach to the active site but instead bind to an allosteric site on the enzyme. This binding alters the shape of the active site so that enzymatic activity is reduced or blocked completely (FIGURE 5.11). Some enzymes have more than one allosteric site, some inhibitory sites and some activation sites (described earlier), allowing the cell to regulate enzyme function precisely. ▶ ANIMATIONS: Substrate Pathway operates Pathway shuts down Enzyme 1 Bound end-product (allosteric inhibitor) Allosteric site Feedback inhibition Intermediate A Enzyme-Substrate Interaction: Noncompetitive Inhibition Cells often control the action of enzymes through feedback inhibition (also called negative feedback or end­product inhibition). Allosteric feedback inhibition functions in much the way a thermostat controls a heater. As the room gets warmer, a sensor inside the thermostat changes shape and sends a signal that turns off the heater. Similarly, in metabolic feedback inhibition, the end-product of a series of reactions is an allosteric inhibitor of an enzyme in an earlier part of the pathway (FIGURE 5.12). Because the product of each reaction in the pathway is the substrate for the next reaction, inhibition of the first enzyme in the series inhibits the entire pathway, thereby saving the cell energy. For example, in Escherichia coli, the presence of the amino acid isoleucine allosterically inhibits the first enzyme in the pathway that produces isoleucine. In this manner, the bacterium prevents the synthesis of isoleucine when this amino acid is already available. When isoleucine is depleted, the pathway is no longer inhibited, and isoleucine production resumes. To this point, we have viewed the concept of metabolism as a collection of chemical reactions (pathways) that can be categorized as either catabolic (breaking down) or anabolic (building up). Because enzymes are required to lower the activation energy of these reactions, we examined them in some detail. Active site Enzyme Distorted active site (enzymatic activity is reduced or stopped) Enzyme 2 Intermediate B End-product Enzyme 3 ▲ FIGURE 5.12 Feedback inhibition. The end-product of a metabolic pathway allosterically inhibits the initial step, shutting down the pathway. Energy is also critical to metabolism, so we examined redox reactions as a means of transferring energy within cells. We saw, for example, that redox reactions and carrier molecules are used to transfer energy from catabolic pathways to ATP, a molecule that stores energy in cells. TELL ME WHY How can oxidation take place in an anaerobic environment, that is, without oxygen? Allosteric site Allosteric inhibitor ▲ FIGURE 5.11 Noncompetitive inhibition at an allosteric site. Inhibition results from a change in the shape of the active site when an inhibitor binds to an allosteric site. M05_BAUM7206_05_SE_C05.indd 132 We will now consider how cells acquire and utilize metabolites, which are used to synthesize the macromolecules necessary for growth and, eventually, reproduction—the ultimate goal of metabolism. We will also consider in more detail the phosphorylation of ADP to make ATP. 11/23/16 3:00 AM       Carbohydrate Catabolism GLYCOLYSIS Carbohydrate Catabolism Respiration LE A R N I N G O U TC O M E 5.8 In general terms, describe the three stages of aerobic glucose catabolism (glycolysis, the Krebs cycle, and an electron transport chain), including their substrates, products, and net energy production. ATP NADH ATP Pyruvic acid (or derivative) 2 Pyruvic acid C O2 Formation of fermentation end-products Acetyl-CoA C O2 FADH2 EL EC TR ON TR AN SP OR T KREBS CYCLE NADH ATP CH AIN (ET C) e– ADP ATP Electrons Final electron acceptor ▲ FIGURE 5.13 Summary of glucose catabolism. Glucose catabolism begins with glycolysis, which forms pyruvic acid and two molecules of both ATP and NADH. Two pathways branch from pyruvic acid: respiration and fermentation. In aerobic respiration (shown here), the Krebs cycle and the electron transport chain completely oxidize pyruvic acid to CO2 and H2O, in the process synthesizing many molecules of ATP. Fermentation results in the incomplete oxidation of pyruvic acid to form organic fermentation products. Which of the blue ovals represents a reduced chemical?  ▶ ANIMATIONS: To see a 3-D animation on metabolism, go to the MasteringMicrobiology Study Area and watch the MicroFlix. ­ Glycolysis4 is commonly the first step in the catabolism of glucose via both respiration and fermentation. Glycolysis in most organisms follows the Embden­Meyerhof­Parnas (EMP) pathway, which is named after the scientists who discovered it. In general, as its name implies, glycolysis involves the splitting of a six-carbon glucose molecule into two threecarbon sugar molecules. When these three-carbon molecules are oxidized to pyruvic acid, some of the energy released is stored in molecules of ATP and NADH. Cells can use many of the intermediate molecules in glycolysis as precursor metabolites. ▶ ANIMATIONS: Glycolysis: Overview The EMP pathway of glycolysis occurs in the cytosol and can be divided into three stages involving a total of 10 steps (FIGURE 5.14). Each step is catalyzed by its own enzyme: M05_BAUM7206_05_SE_C05.indd 133 NADH Figure 5.13 The upper one, with an additional electron, is reduced. Glycolysis 3Cellular respiration is often referred to simply as respiration, which should not be confused with breathing, also called respiration. 4From Greek gleukos, meaning "sweet wine," and lysis, meaning “to loosen.” Fermentation (EMP) Many organisms oxidize carbohydrates as their primary energy source for anabolic reactions. Glucose is used most commonly, though other sugars, amino acids, and fats are also utilized, often by first being converted into glucose. Glucose is catabolized via one of two processes: either via cellular respiration3—a process that results in the complete breakdown of glucose to carbon dioxide and water—or via fermentation, which results in organic waste products. As shown in FIGURE 5.13, both cellular respiration and fermentation begin with glycolysis (glī-kol´i-sis), a process that catabolizes a single molecule of glucose to two molecules of pyruvic acid (also called pyruvate) and results in a small amount of ATP production. Respiration then continues via the Krebs cycle and an electron transport chain, which results in a significant amount of ATP production. Fermentation involves the conversion of pyruvic acid into other organic compounds. Because it lacks the Krebs cycle and an electron transport chain, fermentation results in the production of much less ATP than does respiration. The following is a simplified discussion of glucose catabolism. To help understand the basic reactions in each of the pathways of glucose catabolism, pay special attention to three things: the number of carbon atoms in each of the intermediate products, the relative numbers of ATP molecules produced in each pathway, and the changes in the coenzymes NAD + and FAD as they are reduced and then oxidized back to their original forms. 1. Energy-investment stage (steps 1 – 3 ). As with money, one must invest before a profit can be made. In this case, the Glucose energy in two molecules of ATP is invested to phosphorylate a six-carbon glucose molecule and rearrange its atoms to form fructose 1,6-bisphosphate. 2. Lysis stage (steps 4 and 5 ). Fructose 1,6-bisphosphate is cleaved into glyceraldehyde 3-phosphate (G3P)5 and dihydroxyacetone phosphate (DHAP). Each of these compounds contains three carbon atoms and is freely convertible into the other. DHAP is converted to G3P before the next stage. 5G3P is also known as phosphoglyceraldehyde, or PGAL. 11/23/16 3:00 AM r e 5   Microbial Metabolism hapt       C 3. Energy-conserving stage (steps 6 – 10 ). Each G3P is oxidized to pyruvic acid, yielding two ATP molecules each, for a total of four ATP; it is also oxidized to pyruvic acid, yielding another two ATP molecules, for a total of four ATP molecules. ▶ ANIMATIONS: Glycolysis: Steps GLYCOLYSIS Respiration Fermentation Our study of glycolysis provides our first opportunity to study substrate­level phosphorylation (see steps 1 , 3 , 7 , and 10 ). Let’s examine this important process more closely by considering the 10th and final step of glycolysis. Each of the two phosphoenolpyruvic acid (PEP) molecules produced in step 9 of glycolysis is a three-carbon compound ENERGY-INVESTMENT STAGE C C C C C C Glucose Step 1 . Glucose is phosphorylated by ATP to form glucose 6-phosphate. 1 (EMP) KREBS CYCLE Steps 2 and 3 . The atoms of glucose 6-phosphate are rearranged to form fructose 6-phosphate. Fructose 6-phosphate is phosphorylated by ATP to form fructose 1,6-bisphosphate. EL TR ECTR A O CH NSP N AIN OR T 2 ATP 3 ADP P P C C C C C C Fructose 1,6-bisphosphate Step 4 . Fructose 1,6-bisphosphate is cleaved to form glyceraldehyde 3-phosphate (G3P) and dihydroxyacetone phosphate (DHAP). 4 Step 5 . DHAP is rearranged to form another G3P. Step 6 . Inorganic phosphates are added to the two G3P, and two NAD+ are reduced. P C C C Glyceraldehyde 3-phosphate (G3P) NADH P o id e NADH P ADP 7 ATP 8 P P C C C 1,3-bisphosphoglyceric acid ADP 7 ATP P C C C 3-phosphoglyceric acid 8 H2O 9 H2O 9 P P C C C Phosphoenolpyruvic acid (PEP) C C C Phosphoenolpyruvic acid (PEP) 10 ADP ATP C C C Pyruvic acid 10 ADP ATP C C C Pyruvic acid utor V T NAD+ 6 P C C C 3-phosphoglyceric acid Step 10 . Two ADP are phosphorylated by substrate-level phosphorylation to form two ATP. Two pyruvic acid are formed. P C C C Glyceraldehyde 3-phosphate (G3P) NAD+ P P C C C 1,3-bisphosphoglyceric acid Steps 8 and 9 . The remaining phosphates are moved to the middle carbons. A water molecule is removed from each substrate. P C C C Dihydroxyacetone phosphate (DHAP) 5 6 Step 7 . Two ADP are phosphorylated by substrate-level phosphorylation to form two ATP. ADP P C C C C C C Glucose 6-phosphate LYSIS STAGE ENERGY-CONSERVING STAGE ATP ▲ FIGURE 5.14 Glycolysis by the EMP pathway. Glucose is cleaved and ultimately transformed into two molecules of pyruvic acid in this process. Four ATPs are formed and two ATPs are used, so the result is a net gain of two ATPs. Two molecules of NAD + are reduced to NADH. (For simplification, only carbon atoms and phosphate are shown.) M05_BAUM7206_05_SE_C05.indd 134 11/23/16 3:00 AM       Carbohydrate Catabolism called an electron transport chain, that passes electrons to a chemical not derived from cellular metabolism. Cellular Respiration The Krebs Cycle ­ containing a high-energy phosphate bond. In the presence of a specific holoenzyme (which requires a Mg2+ cofactor), the high-energy phosphate in PEP (one substrate) is transferred to an ADP molecule (a second substrate) to form ATP (step 10 and FIGURE 5.15); the direct transfer of the phosphate between the two substrates is the reason the process is called substrate-level phosphorylation. A variety of substrate-level phosphorylations occur in metabolism. As you might expect, each type has its own enzyme that recognizes both ADP and the enzyme’s specific substrate molecule. In glycolysis, t wo ATP molec ules are i nvested by substrate-level phosphorylation to prime glucose for lysis, and four molecules of ATP are produced by substrate-level phosphorylation. Therefore, a net gain of two ATP molecules occurs for each molecule of glucose that is oxidized to pyruvic acid. Glycolysis also yields two molecules of NADH. Synthesis of Acetyl-CoA Before pyruvic acid can enter the Krebs cycle for respiration, it must first be converted to acetyl­coenzyme A, or acetyl-CoA (as´etilkō-ā; FIGURE 5.16). Enzymes in the pyruvate dehydrogenase complex carry this out, first by removing one carbon from pyruvic acid as CO2—a reaction called a decarboxylation 1 . An enzyme then joins the remaining two-carbon molecule (called acetate) to coenzyme A with a high-energy bond to form acetyl-CoA 2 . This reaction also produces one molecule of NADH 3 . Recall that glycolysis produced two molecules of pyruvic acid from each molecule of glucose. Therefore, this stage produces two molecules of acetyl-CoA, two molecules of CO2, and two molecules of NADH. At this point in the catabolism of a molecule of glucose, a great amount of energy remains in the bonds of acetyl-CoA. The Krebs cycle6 is a “circular” series of eight enzymatically catalyzed reactions that transfer much of this stored energy via electrons to the coenzymes NAD + and FAD (the cycle is diagrammed in FIGURE 5.17). The two carbon atoms in acetate LE A R N I N G O U TC O M E S 5.9 Discuss the roles of acetyl-CoA, the Krebs cycle, and electron transport in carbohydrate catabolism. 5.10 Contrast electron transport in aerobic and anaerobic respiration. 5.11 Identify four classes of carriers in electron transport Respiration phosphorylation of ATP. After glucose has been oxidized via glycolysis or one of the alternative pathways considered shortly, a cell uses the resultant pyruvic acid molecules to complete either cellular respiration or fermentation (which we will discuss in a later section). Our topic here—cellular respiration—is a metabolic process that involves the complete oxidation of substrate molecules and then production of ATP by a series of redox reactions. The three stages of cellular respiration are (1) synthesis of acetyl-CoA, (2) the Krebs cycle, and (3) a final series of redox reactions, GLYCOLYSIS chains. 5.12 Describe the role of chemiosmosis in oxidative O Fermentation H3C O C C OH Pyruvic acid (EMP) C O2 1 O KREBS CYCLE Decarboxylation H H3 C C Acetate EL TR ECTR A O CH NSP N AIN OR T CoA Coenzyme A 2 Phosphoenolpyruvate (PEP) CH2 Holoenzyme 3 CH3 C O C O O H P Mg2+ ADP H+ NAD+ NADH Pyruvic acid O C O C O O H ATP Phosphorylation ▲ FIGURE 5.15 Example of substrate-level phosphorylation. High-energy phosphate bonds are transferred from phosphoenolpyru- CoA H3C C Acetyl-coenzyme A (acetyl-CoA) ▲ FIGURE 5.16 The pyruvate dehydrogenase complex. The enzyme complex acts in a stepwise manner to 1 remove CO2 from pyruvic acid, 2 attach the remaining two-carbon acetate to coenzyme A, and 3 simultaneously reduce a molecule of NAD + to NADH. Because glycolysis produces two molecules of pyruvic acid, two molecules of acetyl-CoA and NADH are produced (only one of each is shown here). E vate (P P) to ATP in this process. What role does Mg2+ play in this reaction? Figure 5.15 Mg2+ is a cofactor of the enzyme. M05_BAUM7206_05_SE_C05.indd 135 6Named for biochemist Hans Krebs, who elucidated its reactions in the 1940s. 11/23/16 3:00 AM r GLYCOLYSIS Respiration e 5   Microbial Metabolism hapt       C Fermentation O (EMP) H3C C CoA Acetyl-CoA KREBS CYCLE EL TR ECTR A O CH NSP N AIN OR T CoA C OOH C C C OOH Oxaloacetic acid NADH NAD+ HOO C C HOO C C C HOO C Citric acid 1 8 C OOH C C C OOH Malic acid 2 C OOH C C C OOH C C OOH Isocitric acid 7 KREBS CYCLE H2O 3 C OOH C C HOO C Fumaric acid NADH + C O2 NAD+ CoA 6 FADH2 FAD NAD+ C OOH C C C OOH Succinic acid C OOH C C C CoA 5 CoA GTP GDP ADP ATP 4 C OOH C C C C OOH c-Ketoglutaric acid NADH + C O2 Succinyl-CoA ▲ FIGURE 5.17 The Krebs cycle. 1 Acetyl-CoA enters the Krebs cycle by joining with oxaloacetic acid to form citric acid, releasing coenzyme A. 2 – 4 Two oxidations and two decarboxylations and the addition of coenzyme A yield succinyl-CoA. 5 Substrate-level phosphorylation produces ATP and again releases coenzyme A. 6 – 8 Further oxidations and rearrangements regenerate oxaloacetic acid, and the cycle can begin anew. Two molecules of acetyl-CoA enter the cycle for each molecule of glucose undergoing glycolysis. M05_BAUM7206_05_SE_C05.indd 136 11/23/16 3:00 AM       Carbohydrate Catabolism are oxidized, and the coenzymes are reduced. The Krebs cycle occurs in the cytosol of prokaryotes and in the matrix of mitochondria in eukaryotes. It is also known as the tricarboxylic acid (TCA) cycle because many of its compounds have three carboxyl ( ¬ COOH) groups. Still another name for the Krebs cycle is the citric acid cycle, for the first compound formed in the cycle. As we will see, cells also use the Krebs cycle for catabolism of lipids and proteins. Anabolism (of citric acid; step 1 ) Isomerization (step 2 ) Redox reactions (steps 3 , 4 , 6 , and Decarboxylations (steps 3 and 4 ) Substrate-level phosphorylation (step Hydration (step 7 ) 8 5 ) ) In the first step of the Krebs cycle, splitting of the highenergy bond between acetate and coenzyme A releases enough energy to enable the binding of the freed two-carbon acetate to a four-carbon compound called oxaloacetic acid, forming the six-carbon compound citric acid (step 1 ). As you study Figure 5.17, notice that after isomerization (step 2 ), the decarboxylations of the Krebs cycle release two molecules of CO2 for each acetyl-CoA that enters (steps 3 and 4 ). Thus, for every pair of carbon atoms that enter the cycle, two are lost to the environment. At this juncture, all six carbon atoms of a molecule of glucose have been lost to the environment: two as CO2 produced during decarboxylation of two molecules of pyruvic acid to form acetyl-CoA, and four as CO2 molecules released in decarboxylations during two turns through the Krebs cycle. (One molecule of acetyl-CoA enters the cycle at a time.) For every two molecules of acetyl-CoA that pass through the Krebs cycle, two molecules of ATP are generated by substrate-level phosphorylation (step 5 ). A molecule of guanosine triphosphate (GTP), which is similar to ATP, can serve as an intermediary in this process. The two molecules of ATP produced by the Krebs cycle are a small amount compared to the energy available. Most of the energy contained in the original glucose molecule is still in electrons, which are carried away by NADH and FADH2. Redox reactions reduce FAD to FADH2 (step 6 ) and NAD + to NADH (steps 3 , 4 , and 8 ); so, for the two molecules of acetyl-CoA derived from the original glucose molecule, six molecules of NADH and two of FADH2 are formed. These coenzymes are the most important molecules of respiration because they carry a large amount of energy that is subsequently used to phosphorylate ADP to ATP. Many of the intermediates of the Krebs cycle are also precursor metabolites; for example, cells can use oxaloacetic acid to make several kinds of amino acids (see Figure 5.29). ▶ ANIMATIONS: Krebs Cycle: Overview, Steps M05_BAUM7206_05_SE_C05.indd 137 ▶ ANIMATIONS: Electron Transport Chain: Overview An electron transport chain consists of a series of membrane-bound carrier molecules that pass electrons from one to another and ultimately to a final electron acceptor. Typically, as we have seen, electrons come from the catabolism of an organic molecule such as glucose; however, microorganisms called lithotrophs (lith´ō-trōfs) acquire electrons from inorganic sources such as H2, NO2 - , or Fe2+ . (Chapter 6 discusses lithotrophs further.) In any case, molecules in the chain pass electrons down the chain to the final acceptor like firefighters of old, who passed buckets of water from one to another until the last one threw the water on a fire. In electron transport, NAD + is an “empty-handed firefighter”; NADH is a “firefighter with a bucket.” As with a bucket brigade, the final step of electron transport is irreversible. Energy from the electrons is used to actively transport (pump) protons (H + ) across the membrane, establishing a proton gradient that generates ATP via a process called chemiosmosis, which we will discuss shortly. To avoid getting lost in the details of electron transport, keep the following critical concepts in mind: • Electrons pass sequentially from one membrane-bound carrier molecule to another, each time losing some energy. Eventually, they pass to a final acceptor molecule. • The electrons’ energy is used to pump protons across the membrane. Electron transport chains are located in the cytoplasmic membranes of prokaryotes (and in the inner mitochondrial membranes of eukaryotes) (Figure 5.18b). Though NADH and FADH2 donate electrons as hydrogen atoms (electrons and protons), many carriers pass only the electrons down the chain. There are four categories of carriers in the transport chains: ­ • • • • • • ­ There are six types of reactions in the Krebs cycle: Electron Transport Scientists estimate that each day an average human synthesizes his or her own weight in ATP molecules and uses them for metabolism, responsiveness, growth, and cell reproduction. ATP turnover in microbes is relatively as copious. In organisms that use respiration, the most significant production of ATP does not occur through glycolysis or the Krebs cycle, but rather through the stepwise release of energy from a series of redox reactions known as an electron transport chain (FIGURE 5.18a). • Flavoproteins are integral membrane proteins, many of which contain flavin, a coenzyme derived from riboflavin (vitamin (vitamin B2).). One form of flavin is flavin mononucleotide (FMN), which is the initial carrier molecule of electron transport chains of mitochondria. The familiar FAD is a coenzyme for other flavoproteins. Like all carrier molecules in an electron transport chain, flavoproteins alternate between reduced and oxidized states. • Ubiquinones (yū-bik´wi-nōns) are lipid-soluble, nonprotein carriers that are so named because they are ubiquitous (present universally) in cells. Ubiquinones are derived from vitamin K. In mitochondria, the ubiquinone is called coenzyme Q. 11/23/16 3:00 AM       CHAPTER 5   Microbial Metabolism GLYCOLYSIS Respiration 35 Fermentation e– 30 (EMP) Path of electrons KREBS CYCLE Approximate energy per electron (kcal/mole) NADH EL TR ECTR A O CH NSP N AIN OR T Prokaryote Exterior Cytoplasmic membrane Cytoplasm Exterior + + + + NADH dehydrogenase and FMN 20 – NADH from glycolysis and Krebs cycle – + 15 NAD+ – – 2 H+ CoQ Reduced Reduced FADH2 Cyt Oxidized ATP 10 2 H+ Cyt Oxidized FAD Reduced Reduced Cyt Final electron acceptor ATP 5 2 H+ Oxidized Dir ect ion 0 e– ele ctr on s fl + + + + + H+ H Cyt b Cyt c e– e– e– FADH2 Oxidized Oxidized e– e– + ow as Reduced the y lo se e– en erg y Cyt a3 FAD + H FADH2 from Krebs cycle – H+ – – + + + H+ + H+ + 4 – – – H+ – – H+ – – Cytoplasm + + + + + + e– e– e– e– Cyt c1 + Exterior H+ H+ – – + + Cyt a – NADH + H+ FeS ATP + e– e– Reduced Reduced + Ubiquinone 1 2 H+ FMN (a) + + H+ e– e– Oxidized NAD+ + H+ 2 Cytoplasmic membrane 25 – 1/ O 2 2 e– e– H+ H+ – – – ATP synthase – – ADP + P 3 (b) H2O ATP ▲ FIGURE 5.18 One possible arrangement of electron transport chain molecules. The exact types and sequences of carrier molecules in electron transport chains vary among organisms. (a) Diagrammatic view of electron and energy transfer. As electrons move down the chain from molecule to molecule (red arrows), energy is released. This energy can be used to synthesize ATP. The ATP icons indicate the approximate point in the chain where enough energy has been released to make an ATP molecule, but the molecules of the chain do not actually synthesize ATP. (b) Electron transport chains are VIDEO located in the cytoplasmic membranes of prokaryotes (as illustrated) and in the inner membranes of mitochondria in eukaryotes. Energy of electrons moving TUTOR down the chain (red arrows) 1 , is used to pump protons (H + ) across the membrane 2 . Eventually, the electrons pass to a final acceptor, in this case, oxygen 3 . Protons then flow through ATP synthase (ATPase), which phosphorylates ADP to make ATP 4 . Approximately one molecule of ATP is generated for every three protons that cross the membrane. Why is it essential that electron carriers of an electron transport chain be membrane bound? Figure 5.18 The carrier molecules of electron transport chains are membrane bound for at least two reasons: (1) Carrier molecules need to be in fairly close proximity so they can pass electrons between each other. (2) The goal of the chain is the pumping of H + across a membrane to create a proton gradient; without a membrane, there could be no gradient. M05_BAUM7206_05_SE_C05.indd 138 11/29/16 8:50 PM       Carbohydrate Catabolism • Metal­containing proteins are a mixed group of integral proteins with a wide-ranging number of iron, sulfur, or copper atoms that can alternate between reduced and oxidized states. Iron-sulfur proteins occur in various places in electron transport chains of many organisms. Copper proteins are found only in electron transport chains involved in photosynthesis (discussed shortly). • Cytochromes (sī´tō-krōms) are integral proteins associated with heme, which is the same iron-containing, nonprotein, pigmented molecule found in the hemoglobin of blood. Iron can alternate between a reduced (Fe2+ ) state and an oxidized (Fe3+ ) state. Researchers named cytochromes with letters and numbers based on the order in which they identified them, so the sequence of carrier molecules can seem out of order; for example, cytochrome b may occur before cytochrome a in an electron transport chain. ANIMATIONS: Electron Transport Chain: Factors Affecting ATP Yield In summary, a number of redox reactions in glycolysis and in the Krebs cycle strip electrons, which carry energy, from glucose molecules and transfer them to molecules of NADH and FADH2. In turn, NADH and FADH2 pass the electrons to electron transport chains. As the electrons move down an electron transport chain, proton pumps use the electrons’ energy to actively transport protons (H + ) across the membrane, creating a proton concentration gradient. Recall, however, that the significance of electron transport is not merely pumping protons across a membrane but the synthesis of ATP. We turn now to the process by which cells synthesize ATP using the proton gradient. ­ ▶ ANIMATIONS: Electron Transport Chain: The Process prokaryotes—particularly archaea called methanogens— reduce carbonate (CO3 2- ) to methane gas (CH4). Medical laboratory scientists can test for products of anaerobic respiration, s u c h a s n i t r i t e, t o h e l p i d e n t i f y s o m e s p e c i e s o f microbes. (Chapter 26 discusses anaerobic respiration as a critical process in recycling nitrogen and sulfur in nature.) ▶ The carrier molecules in electron transport chains are diverse—bacteria typically have different carrier molecules arranged in different sequences from those of archaea or the mitochondria of eukaryotes. Even among bacteria, the makeup of carrier molecules can vary. For example, the pathogens Neisseria (nī-se´rē-ă) and Pseudomonas contain two cytochromes, a and a3—together called cytochrome oxidase—that oxidize cytochrome c; such bacteria are said to be oxidase positive. In contrast, other bacterial pathogens, such as Escherichia, Salmonella (sal´mŏ-nel´ă), and Proteus (prō´tē-ŭs), lack cytochrome oxidase and are thus considered to be oxidase negative. Electrons carried by NADH enter a transport chain earlier than electrons carried by FADH2, which are passed to a ubiquinone. Thus, more energy can be harvested from electrons carried by NADH than from those carried by FADH2 and more molecules of ATP can be generated from NADH than from FADH2. The order and types of electron carriers in electron transport chains vary among organisms. Some bacteria, including E. coli, can even change their carrier molecules to accommodate different environmental conditions. Researchers are not agreed on which carrier molecules are the actual proton pumps or on the number of protons that are pumped as energy from electrons is harvested. Figure 5.18b shows one possibility. In some organisms, the final electron acceptors are oxygen atoms, which, with the addition of hydrogen ions, generate H2O. Because they use oxygen as the final electron acceptor, these organisms are called aerobes and conduct aerobic 7 respiration. Other organisms, called anaerobes,8 perform anaerobic respiration by using inorganic chemicals other than oxygen (or rarely an organic molecule) as the final electron acceptor. The anaerobic bacterium Desulfovibrio (dē´sul-fō-vib´rē-ō), for example, reduces sulfate (SO4 2- ) to hydrogen sulfide gas (H2S), whereas anaerobes in the genera Bacillus (ba-sil´ŭs) and Pseudomonas utilize nitrate (NO3 - ) to produce nitrite ions (NO2 - ), nitrous oxide (N2O), or nitrogen gas (N2). Some 7From Greek aer, meaning “air” (i.e., oxygen), and bios, meaning “life.” M05_BAUM7206_05_SE_C05.indd 139 Chemiosmosis Chemiosmosis is a general term for the use of ion gradients to generate ATP; that is, ATP is synthesized utilizing energy released by the flow of ions down their electrochemical gradient across a membrane. The term should not be confused with osmosis of water. To understand chemiosmosis, we need to review several concepts concerning diffusion and phospholipid membranes (see pp. 68–72 in Chapter 3). Recall that chemicals diffuse from areas of high concentration to areas of low concentration and toward an electrical charge opposite their own. We call the composite of differences in concentration and charge an electrochemical gradient. Chemicals diffuse down their electrochemical gradients. Recall as well that membranes of cells and organelles are impermeable to most chemicals unless a specific protein channel allows their passage across the membrane. A membrane maintains an electrochemical gradient by keeping one or more chemicals in a higher concentration on one side. The membrane’s blockage of diffusion creates potential energy, like water behind a dam. Chemiosmosis uses the potential energy of electrochemical gradients to phosphorylate ADP into ATP. Even though chemiosmosis is a general principle relevant to both oxidative phosphorylation and photophosphorylation, here we consider it as it relates to oxidative phosphorylation. As we have seen, cells use the energy released during the redox reactions of electron transport chains to actively transport protons (H + ) across a membrane. Theoretically, an electron transport chain pumps three pairs of protons for each pair of electrons contributed by NADH, and it pumps two pairs of protons for each electron pair delivered by FADH2. This difference results from the fact that NADH delivers electrons farther up the chain than does FADH2; therefore, energy carried by FADH2 is used to transport one-third fewer protons (see Figure 5.18a). Because lipid bilayers are impermeable to protons, 8The Greek prefix an means “not.” 11/23/16 3:00 AM r e 5   Microbial Metabolism hapt       C the transport of protons to one side of the membrane creates an electrochemical gradient known as a proton gradient, which has potential energy known as a proton motive force. Protons, propelled by the proton motive force, flow down their electrochemical gradient through protein channels, called ATP synthases (ATPases), that phosphorylate molecules of ADP to ATP (see Figure 5.18b 4). Such phosphorylation is called oxidative phosphorylation because the proton gradient is created by the oxidation of carriers in an electron transport chain. In the past, scientists attempted to calculate the exact number of ATP molecules synthesized per pair of electrons that travel down an electron transport chain. However, it is now apparent that phosphorylation and oxidation are not directly coupled. In other words, chemiosmosis does not require exact constant relationships among the number of molecules of NADH and FADH2 reduced, the number of electrons that move down an electron transport chain, and the number of molecules of ATP that are synthesized. Additionally, cells use proton gradients for other cellular processes, including active transport and bacterial flagellar motion, so not every transported electron results in ATP production. Nevertheless, about 34 molecules of ADP per molecule of glucose are oxidatively phosphorylated to ATP via chemiosmosis: three from each of the 10 molecules of NADH generated from glycolysis, the synthesis of acetyl-CoA, and the Krebs cycle; and two from each of the two molecules of FADH2 generated in the Krebs cycle. Given that glycolysis produces a net two molecules of ATP by substrate-level phosphorylation and that the Krebs cycle produces two more, the complete aerobic oxidation of one molecule of glucose by a prokaryote can theoretically yield a net total of 38 molecules of ATP (TABLE 5.3). The theoretical net maximum for eukaryotic cells is generally given as 36 molecules of ATP because energy from two ATP molecules is required to transport NADH (generated by glycolysis) from the cytoplasm into the mitochondria. Metabolic Diversity LE A R N I N G O U TC O M E S 5.13 Compare and contrast the ED and pentose phosphate pathway with EMP glycolysis in terms of energy production and products. 5.14 Describe several examples of the vast metabolic diversity in bacteria. TABLE 5.3 Some bacteria substitute the Entner­Doudoroff (ED) pathway for the EMP pathway. Bacteria using the ED pathway instead of EMP include potentially pathogenic Pseudomonas aeruginosa [soo-dō-mō´nas ā-roo-ji-nōsă] and Enterococcus faecalis [en-terō-kok´s fē-kă´lis]. Some bacteria, such as E. coli, have both ED and EMP pathways. To date, ED has been discovered only in prokaryotes. Whereas EMP generates two ATP, two pyruvate, and two NADH molecules per glucose oxidized, ED produces one ATP, one NADH, and one NADPH molecule. NADPH and NADH both carry high-energy electrons, but NADH is devoted to catabolic pathways and NADPH to anabolic pathways. The pentose phosphate pathway is an alternative to glycolysis for the breakdown of glucose and is named for the phosphorylated pentose (five-carbon) sugars (ribulose, xylulose, and ribose) that it produces. The pathway also yields molecules of nicotinamide adenine dinucleotide phosphate (NADPH) that carries electrons used in anabolic reactions such as photosynthe­ sis (discussed in a later section). Breakdown of glucose by the pentose phosphate pathway nets a single molecule of ATP from each molecule of glucose, so the pathway is half as energy efficient as glycolysis. However, the pathway is essential because the precursor metabolites and NADPH synthesized in the pathway are required to make many important molecules, including DNA nucleotides, steroids, and fatty acids. Fermentation LE A R N I N G O UTC O M E S 5.15 Describe fermentation, and contrast it with respiration. 5.16 List three useful end-products of fermentation, and explain how fermentation reactions are used to identify bacteria. 5.17 Discuss how biochemical tests for metabolic enzymes and products are used in the identification of bacteria. Sometimes cells cannot completely oxidize glucose by cellular respiration. For instance, they may lack sufficient final electron acceptors, as in the case, for example, of a bacterium that lacks oxygen in the anaerobic environment of the colon. Electrons cannot flow down an electron transport chain unless oxidized carrier molecules are available to receive them at the end. Our bucket brigade analogy can help clarify this point. Summary of Ideal Prokaryotic Aerobic Respiration of One Molecule of Glucose Pathway ATP Produced ATP Used Glycolysis 4 2 2 0 Synthesis of acetyl-CoA and Krebs cycle 2 0 8 2 0 0 0 40 2 Net Total 38 E 34 Total lectron transport chain M05_BAUM7206_05_SE_C05.indd 140 NADH Produced FADH 2 Produced 11/23/16 3:00 AM       Carbohydrate Catabolism M05_BAUM7206_05_SE_C05.indd 141 Respiration GLYCOLYSIS Suppose the last person in the brigade did not throw the water but instead held onto two full buckets. What would happen? The entire brigade would soon consist of firefighters holding full buckets of water. An analogous situation occurs in an electron transport chain: All the carrier molecules are forced to remain in their reduced states when there is no final electron acceptor. Without the continual movement of electrons down the chain, protons cannot be transported, the proton motive force is lost, and oxidative phosphorylation of ADP to ATP ceases. Without sufficient ATP, a cell is unable to anabolize, grow, or divide. ATP can be synthesized in glycolysis and the Krebs cycle by substrate-level phosphorylation; after all, together these pathways produce four molecules of ATP per molecule of glucose. However, careful consideration reveals that in respiration, the processes of glycolysis, formation of acetyl-CoA, and the Krebs cycle require a continual supply of oxidized NAD + molecules (see Figures 5.14, 5.16, and 5.17). Electron transport yields the required NAD + , but without a final electron acceptor, this source of NAD + ceases to be available. A cell in such a predicament must use an alternative source of NAD + provided by alternative pathways, called fermentation pathways. In everyday language, fermentation refers to the production of alcohol from sugar, but in microbiology, fermentation has an expanded meaning. Fermentation is the partial oxidation of sugar (or other metabolites) to release energy using an organic molecule from within the cell as the final electron acceptor. In other words, fermentation pathways are metabolic reactions that oxidize NADH to NAD + while reducing cellular organic molecules. Although energetically less efficient than respiration, fermentation has no need for electron acceptors from the environment. FIGURE 5.19 illustrates two common fermentation pathways that reduce pyruvic acid to lactic acid or ethanol, oxidizing NADH in the process. TABLE 5.4 on p. 142 compares fermentation to aerobic and anaerobic respiration with respect to four crucial aspects of these processes. Microorganisms produce a variety of fermentation products depending on the enzymes and substrates available to each. Though fermentation products are wastes to the cells that make them, many are useful to humans, including ethanol (drinking alcohol) and lactic acid (used in the production of cheese, sauerkraut, and pickles) (FIGURE 5.20). Other fermentation products are harmful to human health and industry. For example, fermentation products of the bacterium Clostridium perfringens (klos-trid´ē-ŭm per-frin´jens) are involved in the necrosis (death) of muscle tissue associated with gangrene. Pasteur discovered that bacterial contaminants in grape juice fermented the sugar into unwanted products such as acetic acid and lactic acid, which spoiled the wine (see Chapter 1). Fermentation reactions can be used to identify microbes. For example, Proteus ferments glucose but not lactose, whereas Escherichia and Enterobacter (en´ter-ō-bak´ter) ferment both. Further, glucose fermentation by Escherichia produces mixed Fermentation (EMP) KREBS CYCLE EL TR ECTR A O CH NSP N AIN OR T CH3 O O C C OH Pyruvic acid NADH NAD+ C O2 OH CH3 O CH C Lactic acid O OH CH3 C H Acetaldehyde NADH NAD+ CH3 CH2 OH Ethanol ▲ FIGURE 5.19 Examples of fermentation. In the simplest fer- mentation reaction, NADH reduces pyruvic acid to form lactic acid. Another simple fermentation pathway involves a decarboxylation reaction and reduction to form ethanol. acids (acetic, lactic, succinic, and formic), whereas Enterobacter produces 2,3-butanediol. Common fermentation tests contain a carbohydrate and a pH indicator, which is a molecule that changes color as the pH changes. An organism that utilizes the carbohydrate causes a change in pH, causing the pH indicator to change color. ▶ ANIMATIONS: Fermentation In this section on carbohydrate catabolism, we have spent some time examining glycolysis, alternatives to glycolysis, the Krebs cycle, and electron transport because these pathways are central to metabolism. They generate all of the precursor metabolites and most of the ATP needed for anabolism. We have seen that some ATP is generated in respiration by substrate-level phosphorylation (in both glycolysis and the Krebs cycle) but that most ATP is generated by oxidative phosphorylation via chemiosmosis utilizing the reducing power of NADH and FADH2. We also saw that some microorganisms use fermentation to provide an alternative source of NAD + . 11/23/16 3:00 AM r e 5   Microbial Metabolism hapt       TABLE 5.4 C Comparison of Aerobic Respiration, Anaerobic Respiration, and Fermentation Aerobic Respiration Anaerobic Respiration Fermentation Oxygen Required Yes No No Type of Phosphorylation Substrate-level and oxidative Substrate-level and oxidative Substrate level Final Electron (hydrogen) Acceptor Oxygen NO3 - , SO4 2- , CO3 2- , or externally acquired organic molecules Cellular organic molecules Potential Molecules of ATP Produced per Molecule of Glucose 38 in prokaryotes, 36 in eukaryotes 2–36 2 TELL ME WHY Why do electrons carried by NADH allow for production of 50% more ATP molecules than do electrons carried by FADH2? Thus far, we have concentrated on the catabolism of glucose as a representative carbohydrate, but microorganisms can also use other molecules as energy sources. In the next section, we will examine catabolic pathways that utilize lipids and proteins. Other Catabolic Pathways Lipid and protein molecules contain abundant energy in their chemical bonds (for example, one C16 palmitate fatty acid molecule can generate 129 ATP!) and can also be converted into Glucose NAD+ NADH Pyruvic acid Representive microbes Propionibacterium Aspergillus Lactobacillus Streptococcus Saccharomyces Clostridium NADH CO2, propionic acid Lactic acid CO2, ethanol Acetone, isopropanol NAD+ Swiss cheese Cheddar cheese, yogurt, soy sauce Wine, beer Nail polish remover, rubbing alcohol Fermentation Fermentation products ▲ FIGURE 5.20 Representative fermentation products and the organisms that produce them. All of the organisms are bacteria except Saccharomyces and Aspergillus, which are fungi. Why does Swiss cheese have holes, but cheddar does not? Figure 5.20 Swiss cheese is a product of Propionibacterium, which ferments pyruvic acid to produce CO2; bubbles of this gas cause the holes in the cheese. Cheddar is a product of Lactobacillus, which does not produce gas. M05_BAUM7206_05_SE_C05.indd 142 11/23/16 3:00 AM       Other Catabolic Pathways precursor metabolites. These molecules are first catabolized to produce their constituent monomers, which serve as substrates in glycolysis and the Krebs cycle. C C 3 H2O C C C C C C C C C C C C C C C C C C To step 5 glycolysis C C DHAP C C C C C C C C C C C C C C C C C C C Fatty acid CoA C C C C C C C C C C C C C C C CoA C C C FADH2 To electron transport chain NAD+ C C FAD H2O metabolite production. M05_BAUM7206_05_SE_C05.indd 143 C (a) Hydrolysis 5.19 Explain how proteins are catabolized for energy and is part of the name of this process because enzymes break the bond at the second carbon atom from the end of a fatty acid, and beta is the second letter in the Greek alphabet. Glycerol + Fatty acids C C C C LE A R N I N G O U TC O M E 9Beta C C Protein Catabolism Some microorganisms—notably food-spoilage bacteria, pathogenic bacteria, and fungi—normally catabolize proteins as an important source of energy and metabolites. Most cells catabolize proteins and their constituent amino acids only when carbon sources such as glucose and fat are not available. Generally, proteins are too large to cross cytoplasmic membranes, so prokaryotes typically conduct the first step in the process of protein catabolism outside the cell by secreting proteases (prō´tē-ās-ez)—enzymes that split proteins into their constituent amino acids (FIGURE 5.22). Once released by the action of proteases, amino acids are transported into the cell, where special enzymes split off amino groups in a reaction called deamination. The resulting altered molecules enter the Krebs cycle, and the amino groups are either recycled to synthesize other amino acids or excreted as nitrogenous wastes such as ammonia (NH3) or ammonium ion (NH4 + ). P C C C ADP ATP C C C C C C Lipase olite production. The most common lipids involved in ATP and metabolite production are fats, which typically consist of glycerol and fatty acids (as we saw in Chapter 2). In the first step of triglyceride catabolism, enzymes called lipases hydrolyze the bonds attaching glycerol to the fatty acid chains (FIGURE 5.21a). Subsequent reactions further catabolize the glycerol and fatty acid molecules. Glycerol is converted to DHAP, which, as one of the substrates of glycolysis, is oxidized to pyruvic acid (see Figure 5.14 5 – 10 ). The fatty acids are degraded in a catabolic process known as beta-oxidation9 (FIGURE 5.21b). In this process, enzymes repeatedly split off pairs of the hydrogenated carbon atoms that make up a fatty acid and join each pair to coenzyme A to form acetyl-CoA; this continues until the entire fatty acid has been converted to molecules of acetyl-CoA, which can then feed into the Krebs cycle (see Figure 5.17). Betaoxidation also generates NADH and FADH2, which supply electrons to electron transport chains, ultimately resulting in ATP production. As is the case for the Krebs cycle, the enzymes involved in beta-oxidation are located in the cytosol of prokaryotes and in the mitochondria of eukaryotes. C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C C LE A R N I N G O U TC O M E C C C C Lipid Catabolism 5.18 Explain how lipids are catabolized for energy and metab- Fatty acid chains Glycerol NADH C C C Shorter fatty acid (b) Beta-oxidation C C C C C C CoA C C CoA Acetyl-CoA To Krebs cycle ▲ FIGURE 5.21 Catabolism of a triglyceride molecule. (a) Lipase breaks triglycerides into glycerol and three fatty acids by hydrolysis. Glycerol is converted to DHAP, which can be catabolized via glycolysis and the Krebs cycle. (b) Fatty acids are catabolized via beta-oxidation reactions that produce molecules of acetyl-CoA and reduced coenzymes (NADH and FADH2). TELL ME WHY Why does catabolism of amino acids for energy result in ammonia and other nitrogenous wastes? 11/23/16 3:00 AM … H H O N C C r e 5   Microbial Metabolism hapt       C R R N C C H H O H H O N C C R R N C C H H O H H O N C C R R N C C H H O … Polypeptide Proteases H2O Extracellular fluid H H H O N C C R OH H R Cytoplasmic membrane N C C H H O H O N C C OH Amino acids R ◀ FIGURE 5.22 Protein catabolism. Secreted proteases NH2 2H R C C O O OH To Krebs cycle Thus far, we have examined the catabolism of carbohydrates, lipids, and proteins. Now we turn our attention to the synthesis of these molecules, beginning with the anabolic reactions of photosynthesis. Photosynthesis LE A R N I N G O U TC O M E 5.20 Define photosynthesis. Many organisms use only organic molecules as a source of energy and metabolites, but where do they acquire organic molecules? Ultimately, every food chain begins with anabolic pathways in organisms that synthesize their own organic molecules from inorganic carbon dioxide. Most of these organisms capture light energy from the sun and use it to drive the synthesis of carbohydrates from CO2 and H2O by a process called photosynthesis. Cyanobacteria, purple sulfur bacteria, green sulfur bacteria, green nonsulfur bacteria, purple nonsulfur bacteria, algae, green plants, and a few protozoa are photosynthetic. ▶ ANIMATIONS: Photosynthesis: Overview Chemicals and Structures LE A R N I N G O U TC O M E 5.21 Compare and contrast the basic chemicals and structures involved in photosynthesis in prokaryotes and eukaryotes. M05_BAUM7206_05_SE_C05.indd 144 H Deamination H2O Cytoplasm OH H hydrolyze proteins, releasing amino acids, which are deaminated after uptake to produce molecules used as substrates in the Krebs cycle. R indicates the side group, which varies among amino acids. Photosynthetic organisms capture light energy with pigment molecules, the most important of which are chlorophylls. Chlorophyll molecules are composed of a hydrocarbon tail attached to a light-absorbing active site centered around a magnesium ion (Mg2 + ) (FIGURE 5.23a). The active sites of chlorophyll molecules are structurally similar to the cytochrome molecules found in electron transport chains, except that chlorophylls use Mg2 + rather than Fe2 + . Chlorophylls, typically designated with letters—for example, chlorophyll a, chlorophyll b, and bacteriochlorophyll a—vary slightly in the lengths and structures of their hydrocarbon tails and in the atoms that extend from their active sites. Green plants, algae, photosynthetic protozoa, and cyanobacteria principally use chlorophyll a, whereas green and purple bacteria use bacteriochlorophylls. The slight structural differences among chlorophylls cause them to absorb light of different wavelengths. For example, chlorophyll a from algae best absorbs light with wavelengths of about 425 nm and 660 nm (violet and red), whereas bacteriochlorophyll a from purple bacteria best absorbs light with wavelengths of about 350 nm and 880 nm (ultraviolet and infrared). Because they best use light with differing wavelengths, algae and purple bacteria successfully occupy different ecological niches. Cells arrange numerous molecules of chlorophyll and other pigments within a protein matrix to form light-harvesting matrices called photosystems that are embedded in cellular membranes called thylakoids. Thylakoids of photosynthetic prokaryotes are invaginations of their cytoplasmic membranes (FIGURE 5.23b). The thylakoids of eukaryotes appear to be 11/23/16 3:00 AM       Photosynthesis Thylakoid Photosystem embedded in membrane (sectioned) Chlorophyll Mg Active site Thylakoid membrane Tail (carbon chain) (a) (b) TEM 0.2 µm ▲ FIGURE 5.23 Photosynthetic structures in a prokaryote. (a) Chlorophyll molecules are grouped together in thylakoid membranes to form photosystems. Chlorophyll contains a light-absorbing active site that is connected to a long hydrocarbon tail. The chlorophyll shown here is bacteriochlorophyll a; other chlorophyll types differ in the side chains protruding from the central ring. (b) Prokaryotic thylakoids are infoldings of the cytoplasmic membrane, here in a cyanobacterium. formed from infoldings of the inner membranes of chloroplasts, though thylakoid membranes and the inner membranes are not connected in mature chloroplasts (see Figure 3.41). Thylakoids of chloroplasts are arranged in stacks called grana. An outer chloroplast membrane surrounds the grana, and the space between the outer membrane and the thylakoid membrane is known as the stroma. The thylakoids enclose a narrow, convoluted cavity called the thylakoid space. There are two types of photosystems, named photosystem I (PS I) and photosystem II (PS II), in the order of their discovery. Photosystems absorb light energy and use redox reactions to store this energy in molecules of ATP and nicotinamide adenine dinucleotide phosphate (NADPH). Because they depend on light energy, these reactions of photosynthesis are classified as light-dependent reactions. Photosynthesis also involves light-independent reactions that actually synthesize glucose from carbon dioxide and water. Historically, these reactions were called light and dark reactions, respectively, but this older terminology implies that light-independent reactions occur only in the dark, which is not the case. We first consider the light-dependent reactions of the two photosystems. Light-Dependent Reactions LE A R N I N G O U TC O M E S 5.22 Describe the components and function of the two photosystems, PS I and PS II. 5.23 Contrast cyclic and noncyclic photophosphorylation. M05_BAUM7206_05_SE_C05.indd 145 The pigments of photosystem I absorb light energy and transfer it to a neighboring molecule within the photosystem until the energy eventually arrives at a special chlorophyll molecule called the reaction center chlorophyll (FIGURE 5.24). Light energy from hundreds of such transfers excites electrons in the reaction center chlorophyll, which passes its excited electrons to the reaction center’s electron acceptor, which is the initial carrier of an electron transport chain. As electrons move down this electron transport chain, their energy is used to pump protons across the membrane, creating a proton motive force. In prokaryotes, protons are pumped out of the cell; in eukaryotes, they are pumped from the stroma into the interior of the thylakoids—the thylakoid space. The proton motive force is used in chemiosmosis, in which, you should recall, protons flow down their electrochemical gradient through ATPases, which generate ATP. In photosynthesis, this process is called photophosphorylation, which can be either cyclic or noncyclic. Cyclic Photophosphorylation Electrons moving from one carrier molecule to another in a thylakoid must eventually pass to a final electron acceptor. In cyclic photophosphorylation, which occurs in all photosynthetic organisms, the final electron acceptor is the original reaction center chlorophyll that donated the electrons (FIGURE 5.25a). In other words, when light energy excites electrons in PS I, they pass down an electron transport chain and return to PS I. The energy from the electrons is used to establish a proton gradient that drives the phosphorylation of ADP to ATP by chemiosmosis. ▶ ANIMATIONS: Photosynthesis: Cyclic Photophosphorylation 11/23/16 3:00 AM       Chapter 5   Microbial Metabolism Light Acceptor Reaction center chlorophyll Reaction center To this point, we have examined the use of photosynthetic pigments and thylakoid structure to harvest light energy to produce both ATP and reducing power in the form of NADPH. Next, we examine the light-independent reactions of photosynthesis. ▶ ANIMATIONS: Photosynthesis: Noncyclic Photophosphorylation Light-Independent Reactions LE A R N I N G O UTC O M E S 5.24 Contrast the light-dependent and light-independent reac- tions of photosynthesis. 5.25 Describe the reactants and products of the Calvin-Ben- son cycle. Noncyclic Photophosphorylation 1 Fixation of CO2. An enzyme attaches three molecules of carbon dioxide (three carbon atoms) to three molecules of RuBP (15 carbon atoms), which are then split to form six molecules of 3-phosphoglyceric acid (18 carbon atoms). The rubisco enzyme that catalyzes this reaction may be the most abundant protein on the planet. 2 Reduction. Molecules of NADPH reduce the six molecules of 3-phosphoglyceric acid to form six molecules of glyceraldehyde 3-phosphate (G3P; 18 carbon atoms). These reactions require six molecules of ATP and six molecules of NADPH generated in the light-dependent reactions. ­ Some photosynthetic bacteria and all plants, algae, and photosynthetic protozoa utilize noncyclic photophosphorylation as well. (Exceptions are green and purple sulfur bacteria.) Noncyclic photophosphorylation, which requires both PS I and PS II, generates molecules of ATP and also reduces molecules of coenzyme NADP + to NADPH (FIGURE 5.25b). When light energy excites electrons of PS II, they are passed to PS I through an electron transport chain. (Note that photosystem II occurs first in the pathway and photosystem I second; as mentioned earlier, the photosystems were named in the order in which they were discovered, not the order in which they operate.) PS I further energizes the electrons with additional light energy and transfers them through an electron transport chain to NADP + , which is thereby reduced to NADPH. The hydrogen ions added to NADPH come from the stroma in chloroplasts or from the cytosol in photosynthetic bacteria. NADPH subsequently participates in the synthesis of glucose in the light-independent reactions, which we will examine in the next section. In noncyclic photophosphorylation, a cell must constantly replenish electrons to the reaction center of photosystem II. Oxygenic (oxygen-producing) organisms, such as algae, green plants, and cyanobacteria, derive electrons from the dissociation of H2O (see Figure 5.25b). In these organisms, two molecules of water give up their electrons, producing molecular oxygen (O2) as a waste product of photosynthesis. Anoxygenic photosynthetic bacteria get electrons from inorganic compounds such as H2S, resulting in a nonoxygen waste, such as sulfur. TABLE 5.5 on p. 148 compares photophosphorylation to substrate-level and oxidative phosphorylation. M05_BAUM7206_05_SE_C05.indd 146  ▲ FIGURE 5.24 Reaction center of a photosystem. Light energy absorbed by pigments anywhere in the photosystem is transferred to the reaction center chlorophyll, where it is used to excite electrons for delivery to the reaction center’s electron acceptor.  Photosystem I Light-independent reactions of photosynthesis do not require light directly; instead, they use large quantities of ATP and NADPH generated by the light-dependent reactions. The key reaction of the light-independent pathway of photosynthesis is carbon fixation by the Calvin-Benson cycle,10 which involves the attachment of molecules of CO2 to molecules of a fivecarbon organic compound called ribulose 1,5­bisphosphate (RuBP). RuBP is derived initially from phosphorylation of a precursor metabolite produced by the pentose phosphate pathway. The Calvin-Benson cycle is very endergonic—it requires a great deal of energy. Life on Earth is dependent on carbon fixation by this cycle. This cycle occurs in the cytoplasm of photosynthetic bacteria or the interior stroma of chloroplasts in eukaryotes (see Figure 3.42). The Calvin-Benson cycle is reminiscent of the Krebs cycle in that the substrates of the cycle are regenerated. It is helpful to notice the number of carbon atoms during each part of the three steps of the Calvin-Benson cycle (FIGURE 5.26): Regeneration of RuBP. The cell regenerates three molecules of RuBP (15 carbon atoms) from five molecules of G3P (15 carbon atoms). It uses the remaining molecule of glyceraldehyde 3-phosphate to synthesize glucose by reversing the reactions of glycolysis. In summary, ATP and NADPH from the light-dependent reactions drive the synthesis of glucose from CO2 in the light-independent reactions of the Calvin-Benson cycle. For every three molecules of CO2 that enter the Calvin-Benson 3 10  Possible path of energy transfer Named for the men who elucidated its pathways. 06/03/2018 08:02       Photosynthesis Exterior of prokaryote or thylakoid space of chloroplast H+ H+ H+ H+ H+ H+ Cytochromes H+ H+ H+ H+ H+ H+ H+ H+ H+ H+ Light Fe H+ H+ H+ H+ Photosystem I Membrane of prokaryote or of thylakoid in chloroplast e– Reaction center Cu Cytoplasm of prokaryote or stroma of chloroplast H+ H+ ADP + P ATP synthase ATP (a) Cyclic photophosphorylation H+ H+ H2O H+ Exterior of prokaryote or thylakoid space of chloroplast H+ H+ H+ H2O Light Light H+ Quinone H+ H+ H+ H+ H+ H+ O2 H+ H+ H+ H+ H+ Fe Photosystem I H+ H+ NADPase H+ Membrane of prokaryote or of thylakoid in chloroplast e– e– e– e– e– Cytochromes Photosystem II Reaction center NADP+ Cu Reaction center H+ NADPH H+ Cytoplasm of prokaryote or stroma of Chloroplast (b) Noncyclic photophosphorylation ADP + To Calvin-Benson cycle P ATP synthase ATP ▲ FIGURE 5.25 The light-dependent reactions of photosynthesis: cyclic and noncyclic photophosphorylation. (a) In cyclic photophosphorylation, electrons excited by light striking photosystem I travel down an electron transport chain (red arrows) and then return to the photosystem. (b) In noncyclic photophosphorylation, light striking photosystem II excites electrons that are passed to photosystem I, simultaneously establishing a proton gradient. Light energy collected by photosystem I further excites the electrons, which are used to reduce NADP + to NADPH via an electron transport chain. In oxygenic organisms, new electrons are provided by the lysis of H2O (left). In both types of photophosphorylation, the proton gradient drives the phosphorylation of ADP by ATP synthase (chemiosmosis). M05_BAUM7206_05_SE_C05.indd 147 11/23/16 3:00 AM TABLE 5.5 r e 5   Microbial Metabolism hapt       C A Comparison of the Three Types of Phosphorylation Source of Phosphate Source of Energy Location in Eukaryotic Cell Location in Prokaryotic Cell Substrate-Level Phosphorylation Organic molecule High-energy phosphate bond of donor Cytosol and mitochondrial matrix Cytosol Oxidative Phosphorylation Inorganic phosphate (PO4 3 - ) Proton motive force Inner membrane of mitochondrion Cytoplasmic membrane Photophosphorylation Inorganic phosphate (PO4 3 - ) Proton motive force Thylakoid of chloroplast Thylakoid of cytoplasmic membrane cycle, a molecule of glyceraldehyde 3-phosphate (G3P) leaves. Glycolysis is subsequently reversed to anabolically combine two molecules of G3P to synthesize glucose 6-phosphate. are used, and oxygen is released as a waste product; in aerobic respiration, oxygen serves as the final electron acceptor in the oxidation of glucose to produce carbon dioxide and water. In this section, we examined an essential anabolic pathway for life on Earth—photosynthesis, which produces glucose. Next, we will consider anabolic pathways involved in the synthesis of other organic molecules. ▶ ANIMATIONS: Photosynthesis: Light-Independent Reaction The processes of oxygenic photosynthesis and aerobic respiration complement one another to complete both a carbon cycle and an oxygen cycle. During the synthesis of glucose in oxygenic photosynthesis, water and carbon dioxide 3 C O2 P 6 C C C 3-Phosphoglyceric acid 1 6 P C C C C C 3 P Ribulose bisphosphate (RuBP) 3 ADP 6 ADP P 6 P C C C 1,3-Bisphosphoglyceric acid CALVIN-BENSON CYCLE 3 3 ATP ATP 6 NADPH 2 5 C C C G3P From lightdependent reactions of photosynthesis or catabolic pathways 6 NADP+ P 6 P P 6 C C C Glyceraldehyde 3-phosphate (G3P) 1 C C C G3P P C C C G3P P P C C C C C C Glucose 6-phosphate C C C C C C Glucose M05_BAUM7206_05_SE_C05.indd 148 ◀ FIGURE 5.26 Simplified From the Calvin-Benson cycle or glycolysis diagram of the CalvinBenson cycle. 1 Fixation of CO2. Three molecules of RuBP combine with three molecules of CO2. 2 Reduction. The resulting molecules are reduced to form six molecules of G3P. 3 Generation of RuBP. Five molecules of G3P are converted to three molecules of RuBP, which completes the cycle. Two turns of the cycle also yield two molecules of G3P, which are polymerized to synthesize glucose 6-phosphate. 11/23/16 3:00 AM       Other Anabolic Pathways TELL ME WHY An uninformed student describes the Calvin-Benson cycle as “cellular respiration in reverse.” Why is the student incorrect? Other Anabolic Pathways LE A R N I N G O U TC O M E Anabolic reactions are synthesis reactions. As such, they require energy and a source of precursor metabolites. ATP generated in the catabolic reactions of aerobic respiration, anaerobic respiration, and fermentation and in the initial redox reactions of photosynthesis provides energy for anabolism. Glycolysis, the Krebs cycle, and the pentose phosphate pathway provide 12 basic precursor metabolites from which all macromolecules and cellular structures can be made (TABLE 5.6). Some microorganisms, such as E. coli, can synthesize all 12 precursors, whereas other organisms, such as humans, musvt acquire some precursors in their diets. Many anabolic pathways are the reversal of the catabolic pathways we have discussed; therefore, much of the material that follows has in a sense been discussed in the sections on catabolism. Reactions that can proceed in either direction—toward catabolism or toward anabolism—are said to be amphibolic E a Carbohydrate Biosynthesis LE A R N I N G O UTC O M E 5.26 Define amphibolic reaction. TABLE 5.6 reactions. The following sections discuss the synthesis of carbohydrates, lipids, amino acids, and nucleotides. (Chapter 7 covers anabolic reactions that are closely linked to genetics— the polymerizations of amino acids into proteins and of nucleotides into RNA and DNA.) 5.27 Describe the biosynthesis of carbohydrates. As we have seen, anabolism begins in photosynthetic organisms with carbon fixation by the enzymes of the CalvinBenson cycle to form molecules of G3P. Enzymes use G3P as the starting point for synthesizing sugars, complex polysaccharides such as starch, cellulose for cell walls in algae, and peptidoglycan for cell walls of bacteria. Animals and protozoa synthesize the storage molecule glycogen. Some cells are able to synthesize sugars from noncarbohydrate precursors, such as amino acids, glycerol, and fatty acids, by pathways collectively called gluconeogenesis11 (glū´kō-nē-ōjen´ ĕ-sis; FIGURE 5.27). Most of the reactions of gluconeogenesis are amphibolic, using enzymes of glycolysis in reverse, but four of the reactions require unique enzymes. Gluconeogenesis is highly endergonic and can proceed only if there is an adequate supply of energy. The 12 Precursor Metabolites Examples of Macromolecule Synthesized from Metabolitea Examples of Functional Use Glucose 6-Phosphate Glycolysis Lipopolysaccharide Outer membrane of cell wall Fructose 6-Phosphate Glycolysis Peptidoglycan Cell wall Glyceraldehyde 3-Phosphate (G3P) Glycolysis Glycerol portion of lipids Fats—energy storage Phosphoglyceric Acid Glycolysis Amino acids: cysteine, selenocysteine, glycine, and serine Phosphoenolpyruvic Acid (PEP) Glycolysis Amino acids: phenylalanine, tryptophan, and tyrosine Pyruvic Acid Glycolysis Amino acids: alanine, leucine, and valine Ribose 5-Phosphate Pentose phosphate pathway DNA, RNA, amino acid, and histidine Erythrose 4-Phosphate Pentose phosphate pathway Amino acids: phenylalanine, tryptophan, and tyrosine Acetyl-CoA Krebs cycle Fatty acid portion of lipids a-Ketoglutaric Acid Krebs cycle Amino acids: arginine, glutamic acid, glutamine, and proline Succinyl-CoA Krebs cycle Heme Oxaloacetate Krebs cycle Amino acids: aspartic acid, asparagine, isoleucine, lysine, methionine, and threonine E E E Pathway That Generates the Metabolite nzymes nzymes nzymes E Genome, enzymes nzymes E Cytoplasmic membrane nzymes E Cytochrome electron carrier nzymes xamples given apply to the bacterium E. coli. Greek glukus, meaning “sweet”; neo, meaning “new”; and genesis, meaning “generate.” 11From M05_BAUM7206_05_SE_C05.indd 149 11/23/16 3:00 AM       Chapter 5   Microbial Metabolism Starch, cellulose G L U C O N E O G E N E S I S NADP+ ◀ FIGURE 5.27 The role of gluconeogenesis in the biosynthesis of complex carbohydrates. Complex carbohydrates are synthesized from simple sugar molecules such as glucose, glucose 6-phosphate, and fructose 6-phosphate. Starch and cellulose are found in algae; glycogen is found in animals, protozoa, and fungi; and peptidoglycan is found in bacteria. Glucose 6-phosphate Fructose 6-phosphate Fructose 1,6-bisphosphate Glycerol (from fat) DHAP 2 ADP 2 Oxaloacetic acid ATP ADP C O2 CALVINBENSON CYCLE Peptidoglycan Glucose G3P Amino acids (from protein) Glycogen ATP ATP Pyruvic acid ADP NADPH Fatty acids (from fat) Acetyl-CoA Lipid Biosynthesis Amino Acid Biosynthesis LE A R N I N G O U TC O M E LE A R N I N G O UTC O M E 5.28 Describe the biosynthesis of lipids. 5.29 Describe the biosynthesis of amino acids. ­ Lipids are a diverse group of organic molecules that function as energy storage compounds and as components of membranes (as we saw in Chapter 2). The most energy-efficient energy storage form in cells is triglycerides. The most common form of fat synthesized in cells is phospholipids, to generate membranes. Carotenoids, which are reddish pigments found in many bacterial and plant photosystems, are also lipids. Because of their variety, it is not surprising that lipids are synthesized by a variety of routes. For example, fats are typically synthesized in anabolic reactions that are the reverse of their catabolism; in the case of triglycerides, cells poly merize glycerol and three fatty acids (FIGURE 5.28). Glycerol is derived from G3P generated by the Calvin-Benson cycle and glycolysis; the fatty acids are produced by the linkage of two-carbon acetyl-CoA molecules to one another by a sequence of endergonic reactions that effectively reverse the catabolic reactions of beta-oxidation. Other lipids, such as steroids, are synthesized in complex pathways involving polymerizations and isomerizations of sugar and amino acid metabolites. M05_BAUM7206_05_SE_C05.indd 150 Cells synthesize amino acids from precursor metabolites derived from glycolysis, the Krebs cycle, and the pentose phosphate pathway. Additionally, cells can synthesize many amino acids from other amino acids. Some organisms (such as E. coli and most plants and algae) synthesize all their amino acids from precursor metabolites. Other organisms, including humans, cannot synthesize certain amino acids, called essen­ tial amino acids; these must be acquired in the diet. One extreme example is Lactobacillus (lak´tō-bă-sil´ŭs), a bacterium that ferments milk and produces some cheeses. This microorganism cannot synthesize any amino acids; it acquires all of them by catabolizing proteins in its environment. Precursor metabolites are converted to amino acids by the addition of an amine group. This process is called amination when the amine group comes from ammonia (NH3); an example is the formation of aspartic acid from amination of the Krebs cycle intermediate oxaloacetic acid (FIGURE 5.29a). Amination reactions are the reverse of the catabolic deamination reactions we discussed previously. 06/03/2018 09:46       Other Anabolic Pathways Reverse of beta-oxidation Acetyl-CoA Fatty acids ◀ FIGURE 5.28 Biosynthesis of a triglyceride fat, a lipid. A triglyceride molecule is synthesized from glycerol and three molecules of fatty acid, the precursors of which are produced in glycolysis. Glycolysis Glyceraldehyde 3-phosphate (G3P) CALVINBENSON CYCLE DHAP Phospholipids Glycerol Triglycerides Fats More commonly, however, an amine group is moved from one amino acid to a metabolite, producing a different amino acid. This process is called transamination because an amine group is transferred from one amino acid to form another amino acid (FIGURE 5.29b). All transamination enzymes use a coenzyme, pyridoxal phosphate, which is derived from vitamin B6. Ribozymes of ribosomes polymerize amino acids into proteins. (Chapter 7 examines this energy-demanding process because it is intimately linked with genetics.) are produced from precursor metabolites of glycolysis and the Krebs cycle (FIGURE 5.30): • The five-carbon sugars—ribose in RNA and deoxyribose in DNA—are derived from ribose 5-phosphate from the pentose phosphate pathway. • The phosphate group is derived ultimately from ATP. Purines and pyrimidines are synthesized in a series of ATP-requiring reactions from the amino acids glutamine and aspartic acid derived from Krebs cycle intermediates, ribose 5-phosphate, and folic acid. The latter is synthesized by many bacteria and protozoa but is a vitamin for humans. (Chapter 7 examines the anabolic reactions by which polymerases polymerize nucleotides to form DNA and RNA.) Nucleotide Biosynthesis LE A R N I N G O U TC O M E 5.30 Describe the biosynthesis of nucleotides. TELL ME WHY The building blocks of nucleic acids are nucleotides, each of which consists of a five-carbon sugar, a phosphate group, and a purine or pyrimidine base (see Figure 2.25a). Nucleotides (a) Amination Why is nitrogen required for the production of amino acids by amination? (b) Transamination COOH COOH COOH CH2 CH2 CH2 CH2 NH3 + C O COOH Oxaloacetic acid H C NH2 COOH Aspartic acid H C COOH COOH CH2 NH2 COOH Glutamic acid (an amino acid) + C O COOH Oxaloacetic acid Transaminase with the coenzyme pyridoxal phosphate CH2 COOH CH2 CH2 C O + COOH c-Ketoglutaric acid H C NH2 COOH Aspartic acid (an amino acid) Oxaloacetic acid from Krebs cycle ▲ FIGURE 5.29 Examples of the synthesis of amino acids via amination and transamination. (a) In amination, an amine group from ammonia is added to a precursor metabolite. In this example, oxaloacetic acid is converted into the amino acid aspartic acid. (b) In transamination, the amine group is derived from an existing amino acid. In the example shown here, the amino acid glutamic acid donates its amine group to oxaloacetic acid, which is converted into aspartic acid. Note that transamination is a reversible reaction. M05_BAUM7206_05_SE_C05.indd 151 11/23/16 3:00 AM e 5   Microbial Metabolism r hapt       C ◀ FIGURE 5.30 The biosynthesis DNA and RNA Pyrimidine nucleotides Folic acid (vitamin in humans) PABA of nucleotides. Purine nucleotides Aspartic acid (from Krebs cycle) Glutamine (derived from glutamic acid from Krebs cycle) Photosynthesis Glucose 6-phosphate Glycolysis PENTOSE PHOSPHATE PATHWAY Ribose 5-phosphate Phosphoglyceric acid Glycine Integration and Regulation of Metabolic Functions LE A R N I N G O U TC O M E S 5.31 Describe interrelationships between catabolism and anabolism in terms of ATP and substrates. 5.32 Discuss regulation of metabolic activity. As we have seen, catabolic and anabolic reactions interact with one another in several ways. First, ATP molecules produced by catabolism are used to drive anabolic reactions. Second, catabolic pathways produce precursor metabolites to use as substrates for anabolic reactions. Additionally, most metabolic pathways are amphibolic; they function as part of either catabolism or anabolism as needed. Cells regulate metabolism in a variety of ways to maximize efficiency in growth and reproductive rate. Among the mechanisms involved are the following: ­ • Cells synthesize or degrade channel and transport proteins to increase or decrease the concentration of chemicals in the cytosol or organelles. • Cells often synthesize the enzymes needed to catabolize a particular substrate only when that substrate is available. For instance, the enzymes of beta-oxidation are not produced when there are no fatty acids to catabolize. • If two energy sources are available, cells catabolize the more energy efficient of the two. For example, a bacterium growing in the presence of both glucose and lactose will produce enzymes only for the transport and catabolism of glucose. Once the supply of glucose is depleted, lactoseutilizing proteins are produced. M05_BAUM7206_05_SE_C05.indd 152 • Cells synthesize the metabolites they need, but they typically cease synthesis if a metabolite is available as a nutrient. For instance, bacteria grown in an environment containing an excess of aspartic acid will cease the amination of oxaloacetic acid (see Figure 5.29). • Eukaryotic cells keep metabolic processes from interfering with each other by isolating particular enzymes within membrane-bound organelles. For example, proteases sequestered within lysosomes digest phagocytized proteins without destroying vital proteins in the cytosol. • Cells use inhibitory and excitatory allosteric sites on enzymes to control the activity of enzymes (see Figures 5.9 and 5.11). • Feedback inhibition slows or stops anabolic pathways when the product is in abundance (see Figure 5.12). • Cells regulate catabolic and anabolic pathways that use the same substrate molecules by requiring different coenzymes for each. For instance, NADH is used almost exclusively with catabolic enzymes, whereas NADPH is typically used for anabolism. Note that these regulatory mechanisms are generally of two types: control of gene expression, in which cells control the amount and timing of protein (enzyme) production, and control of metabolic expression, in which cells control the activity of proteins (enzymes) once they have been produced. FIGURE 5.31 diagrams some of the numerous interrelationships among the metabolic pathways discussed in this chapter. ▶ ANIMATIONS: Metabolism:The Big Picture TELL ME WHY Why is feedback inhibition necessary for controlling anabolic pathways? 11/23/16 3:00 AM       Integration and Regulation of Metabolic Functions METABOLIC PATHWAYS FOR THE POLYMERIZATION OF MACROMOLECULES Proteins Nucleic acids Polysaccharides Amino acids Nucleotides Other sugars ATP AND PRECURSOR METABOLIC PATHWAYS GLYCOLYSIS (EMP) PENTOSE PHOSPHATE PATHWAY Lipids GLUCONEOGENESIS Glucose Glucose 6-phosphate Fructose 1,6-bisphosphate Glyceraldehyde 3-phosphate (G3P) Glycerol 3-Phosphoglyceric acid Pyruvic acid Acetyl-CoA Fatty acids KREBS CYCLE NAD+ C O2 NH3 FAD NADH FADH2 ELE CT RO NT ATP ADP CALVINBENSON CYCLE NADP+ RA NS PO RT INTERMEDIATE METABOLIC PATHWAYS CH AIN O2 H2O ADP NADPH KEY: ATP Photosynthetic organisms Catabolic pathway Anabolic pathway Light ▲ FIGURE 5.31 Integration of cellular metabolism (shown in an aerobic organism). Cells possess three major categories of metabolic pathways: E pathways for the polymerization of macromolecules (proteins, nucleic acids, polysaccharides, and lipids), intermediate pathways, and ATP and precursor pathways (glycolysis, Krebs cycle, the pentose phosphate pathway, and the ntner-Doudoroff pathway [not shown]). Cells of photosynthetic organisms also have the Calvin-Benson cycle. M05_BAUM7206_05_SE_C05.indd 153 11/23/16 3:00 AM e 5   Microbial Metabolism r The doctor confirms that the infection has spread to deeper tissues, and he notes areas of necrosis (dead tissue). He tells Arjun and his mom that the fact that the infection hasn’t responded to antibiotics and has spread to deeper tissues is quite serious. The necrotic tissue will need to be surgically removed as soon as possible. During surgery, the dead tissue is removed, and samples are sent to the lab to identify the causative organism. The lab analysis identifies Streptococcus pyogenes in the samples. Intravenous antibiotics are continued post-surgery, and Arjun undergoes a second surgery to remove dead tissue. He remains in the hospital for an additional two weeks and continues oral antibiotics for five weeks after leaving the hospital. With the exception of a few scars, Arjun’s knee ultimately regains full function, and he makes a complete recovery. 1. During Arjun’s infection, S. pyogenes was growing in two very different environments—on the surface of his skin and in the deeper tissue. What types of metabolic pathways would S. pyogenes use in each of these environments, and why? ­ Scraped Knee and Loss of a Leg? ­ hapt       C 2. Why do you think that Arjun must continue to take antibiotics for five weeks following his release from the hospital? Check your answers to Micro in the Clinic Follow-Up questions in the MasteringMicrobiology Study Area. CHAPTER SUMMARY Basic Chemical Reactions Underlying Metabolism (pp. 125–132) 1. Metabolism is the sum of biochemical reactions within the cells of an organism, including catabolism, which breaks down molecules and releases energy, and anabolism, which synthesizes molecules and uses energy. ▶ ANIMATIONS: Metabolism: Overview 2. Precursor metabolites, often produced in catabolic reactions, are used to synthesize all other organic compounds. 3. Reduction reactions are those in which electrons are added to a chemical. A molecule that donates an electron is oxidized. If the electron is part of a hydrogen atom, an oxidation reaction is also called dehydrogenation. Oxidation and reduction reactions always occur in pairs called oxidation-reduction (redox) reactions. ▶ ANIMATIONS: Oxidation-Reduction Reactions 4. Three important electron carrier molecules are nicotinamide adenine dinucleotide (NAD +), nicotinamide adenine dinucleotide (NADP +), and flavin adenine dinucleotide (FAD). 5. Phosphorylation is the addition of phosphate to a molecule. Three types of phosphorylation form ATP: Substrate-level phosphorylation involves the transfer of phosphate from a phosphorylated organic compound to ADP. In oxidative phosphorylation, energy from redox reactions of respiration is used to attach inorganic phosphate (PO4 3 - ) to ADP. Photophosphorylation is the phosphorylation of ADP with inorganic phosphate using energy from light. M05_BAUM7206_05_SE_C05.indd 154 6. Catalysts increase the rates of chemical reactions and are not permanently changed in the process. Enzymes, which are organic catalysts, are often named for their substrates—the molecules on which they act. Enzymes can be classified as hydrolases, isomerases, ligases (polymerases), lyases, oxidoreductases, or transferases, reflecting their mode of action. ▶ ANIMATIONS: Enzymes: Overview ­ This chapter has MicroFlix. Go to the MasteringMicrobiology Study Area to view movie-quality animations for metabolism. 7. Apoenzymes are the portions of enzymes that may require one or more cofactors such as inorganic ions or organic cofactors (also called coenzymes). The combination of an apoenzyme and its cofactors is a holoenzyme. RNA molecules functioning as enzymes are called ribozymes. 8. Activation energy is the amount of energy required to initiate a chemical reaction. 9. Substrates fit into the specifically shaped active sites of the enzymes that catalyze their reactions. Active sites change shape slightly to better fit their substrates—a process described as the induced-fit model. ▶ ANIMATIONS: Enzymes: Steps in a Reaction 10. Enzymes may be denatured by physical and chemical factors such as heat and pH. Denaturation may be reversible or permanent. 11. Enzyme activity proceeds at a rate proportional to the concentration of substrate molecules until all the active sites are filled. 12. Competitive inhibitors block active sites and thereby block enzyme activity. Noncompetitive inhibitors bind to an allosteric site on an enzyme. This binding alters the active site so that it is no longer functional. ▶ ANIMATIONS: Enzymes: Competitive Inhibition; EnzymeSubstrate Interaction: Noncompetitive Inhibition 11/23/16 3:00 AM       Chapter Summary 13. Feedback inhibition (negative feedback) occurs when the final product of a pathway of reactions is an allosteric inhibitor of some previous step in the series. Thus, accumulation of the end product “feeds back” into the pathway a signal that stops the process. Carbohydrate Catabolism (pp. 133–142) 1. Lipids and proteins can be catabolized into smaller molecules, which can be used as substrates for glycolysis and the Krebs cycle. 2. Beta-oxidation is a catabolic process in which enzymes split pairs of hydrogenated carbon atoms from a fatty acid and join them to coenzyme A to form acetyl-CoA. 2. Cellular respiration is a metabolic process that involves the complete oxidation of substrate molecules and the production of ATP following a series of redox reactions. ­ 3. Two carbons from pyruvic acid join coenzyme A to form acetyl-coenzyme A (acetyl-CoA), which then enters the Krebs cycle, a series of eight enzymatic steps that transfer electrons from acetyl-CoA to coenzymes NAD + and FAD. ▶ ANIMATIONS: Krebs Cycle: Overview, Steps o e Photosynthesis (pp. 144–149) 1. Photosynthesis is a process in which light energy is captured by pigment molecules called chlorophylls (bacteriochlorophylls in some bacteria) and transferred to ATP and metabolites. Photosystems are networks of light-absorbing chlorophyll molecules and other pigments held within a protein matrix on membranes called thylakoids. ▶ ANIMATIONS: Photosynthesis: Overview 2. The redox reactions of photosynthesis are classified as lightdependent reactions and light-independent reactions. 3. A reaction center chlorophyll is a special chlorophyll molecule in a photosystem in which electrons are excited by energy harvested from light by many other chlorophyll molecules. The excited electrons of the reaction center are passed to a carrier molecule in an electron transport chain. 4. In cyclic photophosphorylation, the electrons return to the original reaction center after passing down the electron transport chain. ▶ ANIMATIONS: Photosynthesis: Cyclic Photophosphorylation utor 5. The four classes of carrier molecules in electron transport systems are flavoproteins, ubiquinones, metal-containing proteins, and cytochromes. ▶ ANIMATIONS: Electron Transport Chain: The V Process T id 4. An electron transport chain is a series of redox reactions that pass electrons from one membrane-bound carrier to another and then to a final electron acceptor. The energy from these electrons is used to pump protons across the membrane. ▶ ANIMATIONS: Electron Transport Chain: Overview 3. Proteases secreted by microorganisms digest proteins outside the microbes’ cell walls. The resulting amino acids are moved into the cell and used in anabolism or are deaminated and catabolized for energy. ­ o e utor V T id ­ 1. Typical glycolysis involves the splitting a glucose molecule in a three-stage, 10-step process that ultimately results in two molecules of pyruvic acid and a net gain of two ATP and two NADH molecules. ▶ ANIMATIONS: Glycolysis: Overview, Steps Other Catabolic Pathways (pp. 142–144) 6. Aerobes use oxygen atoms as final electron acceptors in their electron transport chains in a process known as aerobic respiration, whereas anaerobes use other inorganic molecules (such as NO3 -, SO4 2 - , and CO23 - or rarely an externally acquired organic molecule as the final electron acceptor in anaerobic respiration. ▶ ANIMATIONS: Electron Transport Chain: Factors Affecting ATP Yield 7. In chemiosmosis, ions flow down their electrochemical gradient across a membrane through ATP synthase (ATPase) to synthesize ATP. ­ 8. A proton gradient is an electrochemical gradient of hydrogen ions across a membrane. It has potential energy known as a proton motive force. 6. In the light-independent pathway of photosynthesis, carbon fixation occurs in the Calvin-Benson cycle, in which CO2 is reduced to produce glucose. ▶ ANIMATIONS: Photosynthesis: Light-Independent Reaction Other Anabolic Pathways (pp. 149–151) 1. Amphibolic reactions are metabolic reactions that are reversible—they can operate catabolically or anabolically. ­ 9. Oxidative phosphorylation and photophosphorylation use chemiosmosis. 5. In noncyclic photophosphorylation, photosystem II works with photosystem I, and the electrons are used to reduce NADP + to NADPH. In oxygenic photosynthesis, cyanobacteria, algae, and green plants replenish electrons to the reaction center by dissociation of H2O molecules, resulting in the release of O2 molecules. Anoxygenic bacteria derive electrons from inorganic compounds such as H2S, producing waste such as sulfur. ▶ ANIMATIONS: Photosynthesis: Noncyclic Photophosphorylation 11. Fermentation is the partial oxidation of sugar to release energy using a cellular organic molecule rather than an electron transport chain as the final electron acceptor. End products of fermentation, which are often useful to humans and aid in laboratory identification of microbes, include acids, alcohols, and gases. ▶ ANIMATIONS: Fermentation M05_BAUM7206_05_SE_C05.indd 155 2. Some cells are able to synthesize glucose from amino acids, glycerol, and fatty acids via a process called gluconeogenesis. ­ 10. The pentose phosphate pathway is an alternative means for the catabolism of glucose that yields fewer ATP molecules than does glycolysis. The pathway produces precursor metabolites and NADPH, which are necessary for synthesis of many cellular chemicals. 3. Amination reactions involve adding an amine group from ammonia to a metabolite to make an amino acid. Transamination occurs when an amine group is transferred from one amino acid to another. 4. Nucleotides are synthesized from precursor metabolites produced by glycolysis, the Krebs cycle, and the pentose phosphate pathway. 11/23/16 3:00 AM 156     Chapter 5   Microbial Metabolism Integration and Regulation of Metabolic Functions (pp. 152–153) 1. Cells regulate metabolism by control of gene expression or metabolic expression. They control the latter in a variety of ways, including synthesizing or degrading channel proteins and enzymes, sequestering reactions in membrane-bound organelles (seen in eukaryotes), and feedback inhibition. ▶ ANIMATIONS: Metabolism: The Big Picture QUESTIONS FOR REVIEW Answers to the Questions for Review (except Short Answer questions) begin on p. A-1. Multiple Choice For each of the phrases in questions 1–7, indicate the type of metabolism referred to, using the following choices: a. anabolism only c. catabolism only b. both anabolism and catabolism (amphibolic) 1. Breaks a large molecule into smaller ones 2. Includes dehydration synthesis reactions 3. Is exergonic 4. Is endergonic 5. Involves the production of cell membrane constituents 6. Includes hydrolytic reactions 7. Includes metabolism 8. Redox reactions          . a. transfer energy b. transfer electrons c. involve oxidation and reduction d. are involved in all of the above 9. A reduced molecule          . a. has gained electrons b. has become more positive in charge c. has lost electrons d. is an electron donor 10. Activation energy          . a. is the amount of energy required during an activity such as flagellar motion b. requires the addition of nutrients in the presence of water c. is lowered by the action of organic catalysts d. results from the movement of molecules 11. Coenzymes are          . a. types of apoenzymes b. proteins c. inorganic cofactors d. organic cofactors 12. Which of the following statements best describes ribozymes? a. Ribozymes are proteins that aid in the production of ribosomes. b. Ribozymes are nucleic acids that produce ribose sugars. c. Ribozymes store enzymes in ribosomes. d. Ribozymes process RNA molecules in eukaryotes. 13. Which of the following does not affect the function of enzymes? a. ubiquinone c. temperature b. substrate concentration d. competitive inhibitors 14. Most oxidation reactions in bacteria involve the          . a. removal of hydrogen ions and electrons b. removal of oxygen M05_BAUM7206_05_SE_C05.indd 156 c. addition of hydrogen ions and electrons d. addition of hydrogen ions 15. Under ideal conditions, the fermentation of one glucose molecule by a bacterium allows a net gain of how many ATP molecules? a. 2 c. 38 b. 4 d. 0 16. Under ideal conditions, the complete aerobic oxidation of one molecule of glucose by a bacterium allows a net gain of how many ATP molecules? a. 2 c. 38 b. 4 d. 0 17. Which of the following statements about the Entner-Doudoroff pathway is false? a. It is a series of reactions that synthesizes glucose. b. Its products are sometimes used to determine the presence of Pseudomonas. c. It is a pathway of chemical reactions that catabolizes glucose. d. It is an alternative pathway to glycolysis. 18. Reactions involved in the light-independent reactions of photosynthesis constitute the          . a. Krebs cycle c. Calvin-Benson cycle b. Entner-Doudoroff d. pentose phosphate pathway pathway 19. The glycolysis pathway is basically          . a. catabolic c. anabolic b. amphibolic d. cyclical 20. A major difference between anaerobic respiration and anaerobic fermentation is          . a. in the use of oxygen b. that the former requires breathing c. that the latter uses organic molecules within the cell as final electron acceptors d. that fermentation only produces alcohol Matching 1.       Occurs when energy from a compound containing phosphate reacts with ADP to form ATP 2.       Involves formation of ATP via reduction of coenzymes in the electron transport chain 3.       Begins with glycolysis 4.       Occurs when all active sites on substrate molecules are filled A. Saturation B. Oxidative phosphorylation C. Substrate-level phosphorylation D. Photophosphorylation E. Carbohydrate catabolism 11/23/16 3:00 AM       Questions for Review Fill in the Blanks           1. The final electron acceptor in cyclic photophosphorylation is .           2. Two ATP molecules are used to initiate glycolysis. Enzymes generate molecules of ATP for each molecule of glucose that undergoes glycolysis. Thus, a net gain of molecules of ATP is produced in glycolysis. 2. Label the diagram below to indicate acetyl-CoA, electron transport chain, FADH2, fermentation, glycolysis, Krebs cycle, NADH, and respiration. Indicate the net number of molecules of ATP that could be synthesized at each stage during bacterial respiration of one molecule of glucose.                     3. The initial catabolism of glucose occurs by glycolysis and/or the and pathways. Glucose 4.           is a cyclic series of eight reactions involved in the catabolism of acetyl-CoA that yields eight molecules of NADH and two molecules of FADH2. .           5. The final electron acceptor in aerobic respiration is ATP ATP C O 7. Anaerobic respiration typically uses (organic/inorganic) molecules as final electron acceptors. Pyruvic acid (or derivative) 2                                         6. Three common inorganic electron acceptors in anaerobic respiration are , , and . 2 Pyruvic acid 8. Complete the following chart: E Category of nzymes End-products Description Catabolizes substrate by adding water C O 2 Isomerase Ligase/polymerase Moves functional groups such as an acetyl group Adds or removes electrons e– ATP Lyase ADP 9. The use of a proton motive force to generate ATP is           Electrons .                     10. The main coenzymes that carry electrons in catabolic pathways are and . VISUALIZE IT! 1. Label the mitochondrion to indicate the location of glycolysis, the Krebs cycle, and electron transport chains. ATP 3. Examine the biosynthetic pathway for the production of the amino acids tryptophan, tyrosine, and phenylalanine in the figure. Where do the initial reactants (erythrose 4-phosphate and PEP) originate? Erythrose 4-phosphate + Phosphoenolpyruvic acid (PEP) Synthetase 1 b Synthetase 2 Synthetase 3 3-Deoxy-D-arabinoheptulosonic acid 7-phosphate (DAHAP) c Feedback inhibition Feedback inhibition Feedback inhibition Tyrosine a M05_BAUM7206_05_SE_C05.indd 157 Phenylalanine Tryptophan 11/23/16 3:00 AM e 5   Microbial Metabolism r hapt       C Short Answer 1. How does amination differ from transamination? 2. Why are enzymes necessary for anabolic reactions to occur in living organisms? 3. How do organisms control the rate of metabolic activities in their cells? 4. How does a noncompetitive inhibitor at a single allosteric site affect a whole pathway of enzymatic reactions? 8. List at least four groups of microorganisms that are photosynthetic. 9. Why do we breathe oxygen and give off carbon dioxide? 10. Why do cyanobacteria and algae take in carbon dioxide and give off oxygen? 11. What happens to the carbon atoms in sugar catabolized by Esche­ richia coli? 12. How do yeast cells make alcohol and cause bread to rise? 5. Explain the mechanism of negative feedback with respect to enzyme action. 13. Where specifically does the most significant production of ATP occur in prokaryotic and eukaryotic cells? 6. Facultative anaerobes can live under either aerobic or anaerobic conditions. What metabolic pathways allow these organisms to continue to harvest energy from sugar molecules in the absence of oxygen? 14. Why are vitamins essential metabolic factors for microbial metabolism? 7. How does oxidation of a molecule occur without oxygen? 15. A laboratory scientist notices that a certain bacterium does not utilize lactose when glucose is available in its environment. Describe a cellular regulatory mechanism that would explain this observation. CRITICAL THINKING 1. Arsenic is a poison that exists in two states in the environment— arsenite (H2AsO3 -) and arsenate (H2AsO4 - ). Arsenite dissolves in water, making the water dangerous to drink. Arsenate is less soluble and binds to minerals, making this form of arsenic less toxic in the environment. Some strains of the bacterium Thermus oxidize arsenic in an aerobic environment but reduce arsenic under anaerobic conditions. In case of arsenic contamination of water, how could scientists use Thermus to remediate the problem? 10. How are photophosphorylation and oxidative phosphorylation similar? How are they different? 2. Explain why an excess of all three of the amino acids mentioned in Visualize It! question 3 (above) is required to inhibit production of DAHAP. 13. Scientists estimate that up to one-third of Earth’s biomass is composed of methanogenic prokaryotes in ocean sediments (Science 295: 2067–2070). Describe the metabolism of these organisms. 3. Why might an organism that uses glycolysis and the Krebs cycle also need the pentose phosphate pathway? 14. A young student was troubled by the idea that a bacterium is able to control its diverse and complex metabolic activities even though it lacks a brain. How would you explain its metabolic control? 4. Describe how bacterial fermentation causes milk to sour. 5. Giardia intestinalis and Entamoeba histolytica are protozoa that live in the colons of mammals and can cause life-threatening diarrhea. Interestingly, these microbes lack mitochondria. What kind of pathway must they have for carbohydrate catabolism? 6. Two cultures of a facultative anaerobe are grown in the same type of medium, but one is exposed to air, and the other is maintained under anaerobic conditions. Which of the two cultures will contain more cells at the end of a week? Why? 7. What is the maximum number of molecules of ATP that can be generated by a bacterium after the complete aerobic oxidation of a fat molecule containing three 12-carbon chains? (Assume that all the available energy released during catabolism goes to ATP production.) 8. In terms of its effects on human metabolism, why is a fever over 40°C often life threatening? 9. Cyanide is a potent poison because it irreversibly blocks cytochrome a3. What effect would its action have on the rest of the electron transport chain? What would be the redox state (reduced or oxidized) of ubiquinone in the presence of cyanide? M05_BAUM7206_05_SE_C05.indd 158 11. Members of the pathogenic bacterial genus Haemophilus require NAD + and heme from their environment. For what purpose does Haemophilus use these growth factors? 12. Compare and contrast aerobic respiration, anaerobic respiration, and fermentation. 15. If a bacterium uses beta-oxidation to catabolize a molecule of the fatty acid arachidic acid, which contains 20 carbon atoms, how many acetyl-CoA molecules will be generated? 16. Some desert rodents rarely have water to drink. How do they get enough water for their cells without drinking it? 17. Why do fatty acids typically contain an even number of carbon atoms? 18. We have examined the total ATP, NADH, and FADH2 production in the Krebs cycle for each molecule of glucose coming through Embden-Meyerhof-Parnas glycolysis. How many of each of these molecules would be produced if the Entner-Doudoroff pathway were used instead of EMP glycolysis? 19. Explain why hyperthermophiles do not cause disease in humans. 20. In addition to extremes in temperature and pH, other chemical and physical agents denature proteins. These agents include ionizing radiation, alcohol, enzymes, and heavy-metal ions. For example, the first antimicrobial drug, arsphenamine, contained the heavy metal arsenic and was used to inhibit the enzymes of the bacterium Treponema pallidum, the causative agent of syphilis. 11/23/16 3:00 AM       Questions for Review Suppose you were able to identify the presence of any specific organic compound. Name a substrate molecule you would find in Pseudomonas and Enterococcus cells but not in human cells. Given that both human and bacterial enzymes are denatured by heavy metals, how was arsphenamine used to treat syphilis without poisoning the patient? Why is syphilis no longer treated with arsenic-containing compounds? 24. Photosynthetic organisms are rarely pathogenic. Why? 21. Figure 5.18b illustrates events in aerobic respiration where oxygen acts as the final electron acceptor to yield water. How would the figure be changed to reflect anaerobic respiration? ­ 25. We have seen that of the two ways ATP is generated via chemiosmosis—photophosphorylation and oxidative phosphorylation—the former can be cyclical, but the latter is never cyclical. Why can’t oxidative phosphorylation be cyclical; that is, why aren’t electrons passed back to the molecules that donated them? ­ 22. Suppose you could insert a tiny pH probe into the space between mitochondrial membranes. Would the pH be above or below 7.0? Why? 23. Even though Pseudomonas aeruginosa and Enterococcus faecalis usually grow harmlessly, they can cause disease. Because these bacteria use the Entner-Doudoroff pathway instead of glycolysis to catabolize glucose, investigators can use clinical tests that provide evidence of the Entner-Doudoroff pathway to identify the presence of these potential pathogens. 26. A scientist moves a green plant grown in sunlight to a room with 24 hours of artificial green light. Will this increase or decrease the plant’s rate of photosynthesis? Why? 27. What class of enzyme is involved in amination reactions? What class of enzyme catalyzes transaminations? CONCEPT MAPPING Using the following terms, fill in the following concept map that describes aerobic respiration. 2 ATP (2) 34 ATP Chemiosmosis Oxidative phosphorylation Substrate-level phosphorylation (2) Electron transport chain Glycolysis Krebs cycle Synthesis of acetyl-CoA Aerobic respiration includes the following 5 processes beginning with 1 2 3 5 4 yields yields yields 6 7 8 produced by produced by produced by 9 10 11 M05_BAUM7206_05_SE_C05.indd 159 06/03/18 4:40 PM