Anabolism Carbohydrates Lecture Notes

OUTLINE: ANABOLISM I: Carbohydrates Introduction and review Transport Glycogenolysis Glycolysis Other sugars Pasteur: Anaerobic vs Aerobic Carbon Assimilation – Calvin Cycle Stage One – Rubisco Carboxylase Know mechanism Oxygenase Glycolate cycle Stage Two – making sugar Stage Three - remaking Ru 1,5P2 Exam-1 material Fermentations Pyruvate Krebs’ Cycle Oxidative Phosphorylation Exam-2 material Electron transport Chemiosmotic theory: Phosphorylation Fat Catabolism Overview and regulation Calvin cycle connections to biosynthesis C4 versus C3 plants Kornberg cycle - glyoxylate Exam-3 material Fatty acid Catabolism Mobilization from tissues (mostly adipose) Activation of fatty acids Transport; carnitine Oxidation: b-oxidation, 4 steps: Protein Catabolism Amino-Acid Degradation Dealing with the nitrogen; Urea Cycle Dealing with the carbon; Seven Families Carbohydrate Biosynthesis in Animals precursors Cori cycle Gluconeogenesis reversible steps irreversible steps – four Nucleic Acid & Nucleotide Degradation energetics 2-steps to PEP in mitochondria: Pyr carboxylase-biotin & PEPCK FBPase G6Pase PHOTOSYNTHESIS: Overview of Photosynthesis Key experiments: Light Reactions Glycogen Synthesis energy in a photon pigments HOW Light absorbing complexes-”red-drop experiment” Reaction center Photosystems (PS) PSII – oxygen from water splitting PSI – NADPH Proton Motive Force – ATP Overview of light reactions UDP-Glc Glycogen synthase branching Pentose-Phosphate Pathway oxidative-NADPH non-oxidative-Ribose 5-P Regulation of Carbohydrate Metabolism Acetyl-CoA/Pyruvate Pyruvate/PEP F6P/FBP: Fru 2,6P2 Glc/Glc6P: sequestration Glycogen: PKA/PP1 Insulin signaling Anaplerotic reactions End of Exam-4 material Regulation of Carbohydrate Metabolism S à protein kinase A No F26P à – insulin glucagon 1 –à Regulation of Carbohydrate Metabolism • Insulin signaling pathway activated when there is a need for energy STORAGE – Causes the translocation of GLUT4 from internal membranes to the plasma membrane. • Glucose enters muscle cells – Increase intracellular [Glc] • In liver, releases glucokinase (liver HK) from binding protein, thus it migrates to the cytosol for activity • In muscle, higher [S] is higher activity Regulation of Carbohydrate Metabolism • Insulin signaling pathway activated when there is a need for energy STORAGE – Causes the translocation of GLUT4 from internal membranes to the plasma membrane. • Glucose enters muscle cells – Increase intracellular [Glc] • In liver, releases glucokinase (liver HK) from binding protein, thus it migrates to the cytosol for activity • In muscle, higher [S] is higher activity – Stimulates Protein Kinase B (PK-B) to phosphorylates GSK-3 at the Ser-4 in the pseudo-substrate site. • This inactivates GSK-3 • Prevents the re-phosphorylation of GS so that it remains ACTIVE S4 2 Regulation of Carbohydrate Metabolism • Insulin signaling pathway activated when there is a need for energy STORAGE – Causes the translocation of GLUT4 from internal membranes to the plasma membrane. • Glucose enters muscle cells – Increase intracellular [Glc] • In liver, releases glucokinase (liver HK) from binding protein, thus it migrates to the cytosol for activity • In muscle, higher [S] is higher activity – Stimulates Protein Kinase B (PK-B) to phosphorylates GSK-3 at the Ser-4 in the pseudo-substrate site. • This inactivates GSK-3 • Prevents the re-phosphorylation of GS so that it remains ACTIVE – Stimulates phosphoprotein phosphatases • Example: ISK phosphorylates the glycogentargeting proteins in muscle (GM) • This dissociates it from PP-1: PP1 is more active Regulation of Carbohydrate Metabolism • Insulin signaling pathway activated when there is a need for energy STORAGE – Causes the translocation of GLUT4 from internal membranes to the plasma membrane. • Glucose enters muscle cells – Increase intracellular [Glc] • In liver, releases glucokinase (liver HK) from binding protein, thus it migrates to the cytosol for activity • In muscle, higher [S] is higher activity – Stimulates Protein Kinase B (PK-B) to phosphorylates GSK-3 at the Ser-4 in the pseudo-substrate site. • This inactivates GSK-3 • Prevents the re-phosphorylation of GS so that it remains ACTIVE – Stimulates phosphoprotein phosphatases • Example: ISK phosphorylates the glycogentargeting proteins in muscle (GM) • This dissociates it from PP-1: PP1 is more active • PP-1 dephosphorylates several proteins – Phosphorylase kinase: inhibits – glycogen phosphorylase: inhibits – Glycogen synthase (GS): activates • GS makes glycogen for energy storage. 3 Regulation of Carbohydrate Metabolism In the Liver: Liver versus the Muscle ↑PFK-2 ↓ FBPase ↑[F26BP] ↑ GK ↑PFK-1 ↓ FBPase ↓ Gluconeogenesis ↑ Gluconeogenesis ↓ HK, PFK-1 ↑FBPase ↑[F6P] ? Muscle does not express PFK2/FBPase-2 Regulation of Carbohydrate Metabolism The Amount of Many Metabolic Enzymes Is Controlled by Transcription TABLE 15-5 Some of the Many Genes Regulated by Insulin Change in gene expression Role in glucose metabolism Increased expression Hexokinase II Hexokinase IV Phosphofructokinase-1 (PFK-1) PFK-2/FBPase-2 Pyruvate kinase Essential for glycolysis, which consumes glucose for energy Glucose 6-phosphate dehydrogenase 6-Phosphogluconate dehydrogenase Malic enzyme Produce NADPH, which is essential for conversion of glucose to lipids ATP-citrate lyase Pyruvate dehydrogenase Produce acetyl-CoA, which is essential for conversion of glucose to lipids Acetyl-CoA carboxylase Fatty acid synthase complex Stearoyl-CoA dehydrogenase Acyl-CoA–glycerol transferases Essential for conversion of glucose to lipids Decreased expression PEP carboxykinase Glucose 6-phosphatase (catalytic subunit) Essential for glucose production by gluconeogenesis 4 Regulation of Carbohydrate Metabolism The Carbohydrate Response Element Binding Protein (ChREBP) Activates Transcription in Response to Glucose Mice with KO of the ChREBP have interesting phenotype. They are intolerant to fructose Anaplerotic Reactions Catabolism Anabolism • We introduced the citric acid cycle as a key catabolic pathway. • It has an equal, if not more important, role in anabolism. – The biosynthesis of biological precursors has to begin with elementary materials – Many, if not most, of these starting points come from the several intermediates in the Kreb’s cycle. – This was first appreciated by Hans Kornberg: how to organisms grow on carbohydrates only? o The term comes from the Greek, to ”fill up” or replenish that without this replenishment, the TCA cycle would grind to a halt o Anaplerotic reactions are critical o Recall Enzyme Kornberg discovered (PEPCK) 5 Anaplerotic Reactions Kornberg Cycle = Glyoxylate Cycle • Was intrigued by the fact that bacteria could grow very effectively on a little ammonium and phosphate salts with acetate (2C) • From these they can synthesize all the components of the cell; DNA, DNA, proteins, membrane lipids, cytochromes, everything….. • How do you build all this from a 2-carbon compound knowing how the Kreb’s cycle works? - 2 Acetyl-CoA + NAD+ ⇌ Succinate + 2 CoASH + NADH 6 Anaplerotic Reactions • Purine Nucleotide Cycle – was first thought be be part of nucleotide degradation or synthesis – In muscle, its now realized as an important anaplerotic pathway Myoadenylate deaminase deficiency Loss of AMP deaminase-1, the muscle-specific isozyme Causes exercise-induced muscle pain H2O + Aspartate + GTP ⇌ NH4+ + GDP + Pi + Fumarate Anaplerotic Reactions • Intermediates in the citric acid cycle can be used in biosynthetic pathways. • Must replenish the intermediates in order for the cycle and central metabolic pathway to continue. • In animals, these 4-carbon intermediates are formed by carboxylation of 3-carbon precursors. Aspartate + GTP AMP deaminase, adenylosuccinate Pyruvate carboxylase deficiency – an inherited metabolic disorder where anaplerosis is greatly reduced. – What is the problem? – How to treat this disorder? – Other anaplerotic substrates such as the odd-carbon-containing triglyceride triheptanoin are used NH4+ + GDP + Pi + Fumarate synthetase, adenylosuccinate lyase Enzyme Kornberg discovered Muscle 7 ANABOLISM I: Summary What we learned: • Gluconeogenesis, a process by which cells can use a variety of metabolites for the synthesis of glucose • The differences between glycolysis and gluconeogenesis – how they are both made thermodynamically favorable – how they are differentially regulated to avoid a futile cycle • The pentose phosphate pathway, a process by which cells can generate pentose phosphates and NADPH. The pentose phosphates can be regenerated into glucose 6-phosphate, for which NO ATP is required. • living organisms regulate the flux of metabolites through metabolic pathways by: – increasing or decreasing enzyme concentrations – activating or inactivating key enzymes in the pathway • the activity of key enzymes in glycolysis and gluconeogenesis is tightly and coordinately regulated via various activating and inhibiting metabolites (Fru 2,6P2) • glycogen synthesis and degradation is regulated by hormones insulin, epinephrine, and glucagon that report on the levels of glucose in the body • the citric acid cycle plays important anabolic roles in the cell: Anaplerosis • organisms have multiple ways to replenish intermediates that are used in other pathways: Lipid and Nitrogen biosynthesis…....... End of Material for Exam 4 8 ANABOLISM II: Biosynthesis of Fatty Acids & Lipids ANABOLISM II: Biosynthesis of Fatty Acids & Lipids 1. Biosynthesis of fatty acids 2. Regulation of fatty acid degradation and synthesis 3. Assembly of fatty acids into triacylglycerol and phospholipids 4. Metabolism of isoprenes a. Ketone bodies and Isoprene biosynthesis b. Isoprene polymerization i. Cholesterol ii. iii. Steroids & other molecules Regulation iv. Role of cholesterol in human disease 9 ANABOLISM II: Biosynthesis of Fatty Acids & Lipids Lipid Biosynthesis Fatty Acid Synthesis Fat Catabolism Fatty Acid Degradation Ketone body Utilization Isoprene Biosynthesis Fatty Acid Biosynthesis Catabolism Anabolism • Contrast with Sugars – Lipids have hydro-carbons not carbo-hydrates – more reduced=more energy – Long-term storage vs short-term storage – Lipids are essential for structure in ALL organisms: membrane phospholipids • Catabolism of fatty acids –produces acetyl-CoA –produces reducing power (NADH, FADH2) –takes place in the mitochondria • Anabolism of fatty acids –requires acetyl-CoA and sufficient carbohydrates –requires reducing power from NADPH –takes place in cytosol in animals, chloroplast in plants Anabolism of fat will occur only when there is excess, carbon, electrons, and DG Acetyl-CoA & carbohydrates How is this monitored in the cell? NADPH ATP 10