Catabolism: Network of chemical reactionsthat degrade organic nutrients into simple(lower) energy end products in order to extract chemical energy that can be used by the cell for useful purposes.Anabolism: Network of reactions thatconvert small simple (lower energy)Molecules to increasing more complexand larger molecules.Glucose CO2+ Water+ Useful Energy (ATP)Enzymes (Chapter 6)Chemistry 153A-18/17/06Handout #3ENZYMESThe individual steps in the anabolic and catabolic reactions are catalyzed by proteins (enzymes)Enzymes: Biological catalysts. Almost always proteins.(1) Increase rates of reactions(2) Catalyze reactions at physiological conditions (mild)(3) Have a high degree of specificity (e.g. only A is converted to B)(4) Can be regulated (e.g. A is only converted to B under certain conditions)Properties of EnzymesReview of Basic Reaction KineticsKinetics: study of the rates at which chemicalreactions occurSPUncatalyzed Unimolecular (first order)kfRate constantFirst orderUncatalyzed Bimolecular (Second order)A + B Pk2Review of Reaction Kinetics (and thermodynamics)SPkfkbActivation energy: Free energy difference between ground state and transition statekfPlanck’s constantBoltzmann’s constantThermodymamicsKineticsUnimolecular reactionEnzyme catalyzed reactions are much faster than the corresponding uncatalyzed reactionE + S ES EP E + PESEnzyme catalyzed rate / non-catalyzed rate•Enzymes are biological catalysts•The vast majority of enzymes are proteinsEnzyme NomenclatureSome enzymes need cofactors to work. Chemical component that helps mediate the chemistry of the catalyzed reaction.•Metal cofactors=Holoenzyme Apoenzyme (apoprotein)+Cofactor•Metallorganic or organic cofactors(also called coenzymes)Types of CofactorsOverview of how enzymes increase the rates of reaction(1) Enzymes stabilize the transition state (ES ) relative to the ES complex(2) Enzymes reduce the entropic cost associated with the reaction (entropy reduction)(3) General Acid-Base Catalysis. (4) Metal Ion Catalysis: Metal ion in the active site assists in the enzymatic catalysis (~1/3 of all enzymes)(5) Covalent Catalysis: transient covalent bond between enzyme and substrate is part of the reaction pathway.E + S ES EP E + PThe same set of non-covalentinteractions that enable a proteinto fold are involved in stabilizingthe interaction between the substrate and enzyme.The enzyme provides a complementarybinding site optimized for the substrateEnzyme Active SiteESInduced fit mechanism is most prevalent in enzymes. The enzyme active site adapts its structure to interact with the substrate and transition states.Enzyme Active SiteHow enzymes increase reaction rates(1) Enzymes stabilize the transition state (ES ) relative to the ES complexE + S ES E + PESEP(2) Enzymes reduce the entropic cost associated with the reaction (entropy reduction)A + B Æ PenzymeA BPEffects of proximity on non-enzymatically catalyzed reactions(3) General Acid-Base Catalysis.Amino Acids involved in Acid-base Catalysis(3) General Acid-Base Catalysis. ( a specific example) Ribonuclease A catalyzed hydrolysis of RNA(4) Metal Ion Catalysis: Metal ion in the active site assists in the enzymatic catalysis (~1/3 of all enzymesExample: Carbonic AnhydraseExample: Carbonic Anhydrase(5) Covalent Catalysis: transient covalent bond between enzyme and substrate is part of the reaction pathway. Example: Peptide bond hydrolysis by the serine proteasesThe remainder ofthe reactionMore on this later…Types of EnzymesSerine proteaseH2O( example of a Hydrolase)A Prototypical Enzyme: Serine ProteasesThe Serine Protease Active Site (Catalytic Triad)Serine Protease Mechanism (another representation)Tetetrahedral transition state stabilizationESESMany Serine Proteases that have different tertiary structures, but the nonetheless contain a similar active site (catalytic triad)Convergent EvolutionTo avoid digestion of undesired tissue (molecules) many proteases aresynthesized as an inactive precursor protein called a zymogenSecreted from pancreasZymogenActive ProteaseChymotrypsinogenChymotrypsinZymogens:Small intestineChymotrypsinogen is inactive becauseof an improperly formed active site:(1) Bad Oxyanion hole(2) Substrate binding siteHow chymotrypsinogen is activated(1) Increase rates of reactions(2) Catalyze reactions at physiological conditions (mild)(3) Have a high degree of specificity (e.g. only A is converted to B)(4) Can be regulated (e.g. A is only converted to B under certain conditions)Properties of EnzymesTypes of Substrate Specificity(1) Geometric Specificity(2) Stereospecificity(1) Geometric SpecificityMore on this laterExample Geometric Specificity: Serine protease specificity depends on the “binding-pocket” used to interact with the R1 side chain of its substrateSerine proteaseH2O(2) StereospecificityEnzyme Kinetics (Michaelis-Menten )The Steady-State kineticsExperiment:(1) Mix enzyme + substrate(2) Record rate of product formation as a function ofof time (the velocity of reaction)(3) Plot velocity versus substrate concentration.Goal: Gain insights into the mechanism by studying the kinetics of the reactionThe Steady-State Kinetics-- Choose conditions such that [S] >> [E]-- No back reaction occurs (initial velocity)Michaelis Constant (Km)It can be a measure of the dissociationconstant for the ES complexkcat: the rate constant for any enzyme-catalyzed reactionoperating at saturating substrate concentrations.For reaction with a single rate limiting step kcat is equal torate constant of the rate limiting step.E + S ES E + Pk1k-1k2kcatfor a Michaelis-Menten type reactionCatalytic Efficiency (specificity constant): The rate constant for the enzyme catalyzed conversion of substrate to productat limiting amounts of substrate.Vo ~ Vmax * [S] / KmAs good as you can get…E + S Æ Products d[Products]/dt == 108 to 10 9M-1s-1(diffusion limited)Lineweaver-Burk Plot(reciprocal plot the Michaelis-Menten equation that enables easy extraction of Km and Vmax)Lineweaver-Burk Plot (1/Voversus 1/[S])Feedback Inhibition of Pathway•Enzymes catalyze the individual steps in a metabolic pathway.•The activity of specific enzymes in the pathway are regulated so as tocontrol the flux of metabolites as cellular conditions changeThe activity of enzymes are regulatedRegulatory Enzymes: exhibit decreased or increasedactivity in response to certain signals.Types:Allosteric regulationReversible covalent modificationAllosteric regulation:
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