University:
Glendale Community College
Course:
BIO160 | Introduction to Human Anatomy and Physiology
Academic year:

2020
Views:

252

Pages:

14
Author:

Deandre Simmons

Section 6.3: Enzyme Kinetics §Enzyme kinetics - quantitative study of enzyme catalysis; affinity for substrates and inhibitors §Velocity (rate) - change of a concentration of reactant or product per unit of time §Initial velocity (v0) –beginning when concentration of S greatly exceeds E concentration, no P For reaction: A+BàP § Rate of reaction is given by rate equation § Rate of reaction at a given time rate = K[A]f[B]g § k is rate constant - a proportionality constant § f,g reaction order for reactant § [A] – molar concentration Section 6.3: Enzyme Kinetics §Order of reaction: sum of exponents in the rate equation §Determined experimentally §First order is unimolecular (no collisions required) (A à P) Rate = k[A]1 §Second order is bimolecular (A + B à P) Rate = k[A]1[B]1 Figure 6.3b Enzyme Kinetic Studies §Zero order – reactant independent; enzyme saturated §Half-life -time for one-half of reactant molecules to be consumed Section 6.3: Enzyme Kinetics §Michaelis-Menten Kinetics §Simple reaction SàP §Mechanism for an enzyme-catalyzed reaction k1 k2 E+S k-1 ES E+P §Michaelis constant Km k-1 + k2 Km = k1 §Lower the value of Km, greater affinity of the enzyme for ES complex formation Section 6.3: Enzyme Kinetics §Vmax - maximum velocity, E is saturated Vmax[S] ν0 = [S] + Km Michaelis-Menten Equation §Turnover number, kcat - number substrate molecules converted to product per unit time §Kcat = Vmax /[Et] Figure 6.4 Initial Reaction Velocity v0 and Substrate Concentration [S] for a Typical Enzyme-Catalyzed Reaction §Specificity constant, (kcat/Km) – relationship between catalytic rate & substrate binding affinity Hi specificity constant = low Km & hi Kcat Section 6.3: Enzyme Kinetics §Lineweaver-Burke plot has the form y = mx + b, formula for a straight line Figure 6.6 Lineweaver-Burk or Double-Reciprocal Plot §Slope of the line Km/Vmax §1/Vmax is the Y intercept §-1/Km is the X intercept Section 6.3: Enzyme Kinetics §Multisubstrate Reactions – 2 or more substrates §Bisubstrate reactions §Sequential—reaction cannot proceed until all substrates are bound to the enzyme active site §Ordered – 1st S must bind before 2nd §Random – substrates can bind in any order; product released in any order §Double-Displacement Reactions—first product is released before second substrate binds §Enzyme is altered by first phase of the reaction Section 6.3: Enzyme Kinetics §Inhibitors – substances that reduce enzyme activity §Drugs, antibiotics, food preservatives, poisons, metabolites of biochemical processes §Importance of inhibition §Metabolic pathway regulation §Clinical therapies are based on enzyme inhibition §Functional properties & physical/chemical architecture Section 6.3: Enzyme Kinetics §Reversible inhibitor: substance binds causing inhibition, can be released §Competitive inhibitor: binds to the active (catalytic) site; blocks access §Noncompetitive inhibitor: binds to a site other than the active site; inhibits by changing conformation §Uncompetitive – binding site created after S is bound §Irreversible inhibitor: substance causes permanent impairment §usually involves formation or breaking of covalent bonds to or on the enzyme Section 6.3: Enzyme Kinetics Figure 6.8 Michaelis-Menten Plot of Uninhibited Enzyme Activity Versus Competitive Inhibition §Competitive Inhibitors bind reversibly to the enzyme at active site §Forms enzyme-inhibitor (EI) complex §Increasing S concentration overcomes competitive inhibition ü Increases rate §Vmax unchanged; KmI increases Section 6.3: Enzyme Kinetics §Noncompetitive Inhibitors bind reversibly at a site other than active site Figure 6.10 MichaelisMenten Plot of Uninhibited Enzyme Activity Versus Noncompetitive Inhibition §Changes enzyme conformation §Increased substrate concentration partially reverses inhibition §Vmax reduced; Km unaffected Section 6.3: Enzyme Kinetics §Competitive inhibition increases Km, not Vmax Section 6.3: Enzyme Kinetics §Pure noncompetitive Vmax lowered Km unchanged Section 6.3: Enzyme Kinetics Allosteric enzyme: an oligomer whose biological activity is affected by other substances binding to it §Substances change the enzyme’s activity by altering the conformation(s) of its 3°4°structure Allosteric effector: a substance that modifies the behavior of an allosteric enzyme Figure 6.13 The Kinetic Profile of an Allosteric Enzyme §allosteric inhibitor – excess product §allosteric activator – need product Section 6.3: Enzyme Kinetics §Enzyme Kinetics, Metabolism §Ultimate goal is understanding enzyme kinetics in living organisms §In vitro work does not always reflect in vivo reality §Macromolecular crowding – cell interior highly heterogeneous; influences reaction rates and equilibrium constants in a non-linear way §Systems biologists are using computer modeling, in vitro, and in vivo data to overcome issues