TAILIEUCHUNG - Lecture Principles of biochemistry - Chapter 6: Mechanisms of enzyme action
In this chapter, you will be able to understand: Define transition state, understand basic properties of a transition state, understand why enzymes bind the transition state with higher affinity than the substrate, know common enzymatic mechanisms, know types of amino acid residues that function in enzyme active sites, know what types of RXNS that each participates,. | Chapter 6 Mechanisms of Enzyme Action Activation Energy (AE) – The energy require to reach transition state from ground state. AE barrier must be exceeded for rxn to proceed. Lower AE barrier, the more stable the transition state (TS) The higher [TS], the move likely the rxn will proceed. Enzymatic Catalysis S Ts P Enzymatic Catalysis Transition (TS) State Intermediate Transition state = unstable high-energy intermediate Rate of rxn depends on the frequency at which reactants collide and form the TS Reactants must be in the correct orientation and collide with sufficient energy to form TS Bonds are in the process of being formed and broken in TS Short lived (10–14 to 10-13 secs) Intermediates Intermediates are stable. In rxns w/ intermediates, 2 TS’s are involved. The slowest step (rate determining) has the highest AE barrier. Formation of intermediate is the slowest step. Enzyme binding of substrates decrease activation energy by increasing the initial ground state (brings reactants into correct orientation, decrease entropy) Need to stabilize TS to lower activation energy barrier. ES complex must not be too stable Raising the energy of ES will increase the catalyzed rate This is accomplished by loss of entropy due to formation of ES and destabilization of ES by strain distortion desolvation Transition State Stabilization Transition state analog Equilibrium between ES TS, enzyme drives equilibrium towards TS Enzyme binds more tightly to TS than substrate Mechanistic Strategies Polar AA Residues in Active Sites Common types of enzymatic mechanisms Substitutions rxns Bond cleavage rxns Redox rxns Acid base catalysis Covalent catalysis Substitution Rxns Nucleophillic Substitution– Direct Substitution transition state Nucleophillic = e- rich Electrophillic = e- poor Oxidation reduction (Redox) Rxns Loose e- = oxidation (LEO) Gain e- = reduction (GER) Central to energy production If something oxidized something must be reduced (reducing agent donates e- to oxidizing agent) Oxidations = removal of hydrogen or addition of oxygen or removal of e- In biological systems reducing agent is usually a co-factor (NADH of NADPH) Heterolytic vs homolytic cleavage Carbanion formation (retains both e-) R3-C-H R3-C:- + H+ Carbocation formation (lose both e-) R3-C-H R3-C+ + H:- Free radical formation (lose single e-) R1-O-O-R2 R1-O* + *O-R2 Cleavage Rxns Hydride ion Accelerates rxn by catalytic transfer of a proton Involves AA residues that can accept a proton Can remove proton from –OH, -NH, -CH, or –XH Creates a strong nucleophillic reactant (. X:-) Acid-Base Catalysis : : Acid-Base Catalysis carbanion intermediate Covalent Catalysis 20% of all enzymes employ covalent catalysis A-X + B + E BX + E + A A group from a substrate binds covalently to enzyme (A-X + E A + X-E) The intermediate enzyme substrate complex (A-X) then donates the group (X) to a second substrate (B) (B + X-E B-X + E) Covalent Catalysis Protein Kinases ATP + E + Protein ADP + E + Protein-P A-P-P-P(ATP) + E-OH A-P-P (ADP) + E-O-PO4- E-O-PO4- + Protein-OH E + Protein-O- PO4- The Serine Proteases Trypsin, chymotrypsin, elastase, thrombin, subtilisin, plasmin, TPA All involve a serine in catalysis - thus the name Ser is part of a "catalytic triad" of Ser, His, Asp (show over head) Serine proteases are homologous, but locations of the three crucial residues differ somewhat Substrate specificity determined by binding pocket Serine Proteases are structurally Similar Chymotrpsin Trypsin Elastase Substrate binding specificity
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