BC351 Lecture 5 Protein Function Enzymes Terms Transition state Enzyme substrate complex Catalysis Cofactor Coenzyme Activation energy Substrate Active site Reaction intermediate Covalent Catalysis Acid base catalysis Metal ion catalysis Enzyme Principles 1 The thermodynamic basis of enzymatic catalysis 2 The molecular basis of enzymatic catalysis 3 The interplay between principles 1 and 2 a Use enolase and lysozyme as our specific example I What are enzymes pgs 183 185 a Define the terms i Definition of an enzyme 1 ii Definition of catalysis 1 iii What are enzymes made of 1 What are cofactors and coenzymes a Definition of cofactors and coenzymes i ii II Chemical reactions pgs 186 188 a What determines the rate of a chemical reaction LN05 1 i Peptide bond hydrolysis 1 The reaction a Like all chemical reactions this reaction results in a redistribution of electrons i The peptide bond is broken a planar bond is loss and water is split 2 Does the equilibrium constant determine the rate a The G for peptide hydrolysis is 8kJ mol G R C i From this one might expect the rate to be fairly fast assuming that the Keq does indeed determine rate ii But does it LN05 2 3 So what does determine the rate a Transition state theory O AA1 AA2 OH2 R C N R H AA1 AA2 H2O O H H G AA1 OH AA2 H O R C R C N O H H H i Definition of the transition state intermediate 1 2 Another way to think about it is that this it is a highly unstable state of the molecules as they redistribute their electrons ii What is the likely transition state of peptide hydrolysis 1 The formation of this is unfavorable as a result it s free energy is high and results in an energy barrier between the reactants and products LN05 3 R b Activation energy G or G uncat i Definition of activation energy 1 ii Rate is dependent upon this value 1 If it is small then the rate will be G large 2 If it is large then the rate will be small R C 3 So the peptide bond is kinetically stable b What can a catalyst do i First what CAN T they do 1 Change the equilibrium constant ii Okay what can they do 1 Lower the activation energy a G cat G G R C R C LN05 4 III How do enzymes lower activation energy pgs 188 191 192 194 a Defining terms and building a framework i Substrate 1 ii Active site 1 iii A general enzymatic reaction scheme 1 They can be very simple to very complex E S k1 ES k 1 k2 E P k 2 iv And yet another term 1 Enzyme substrate complex a b Back to the question i Two ways 1 Transition state stabilization 2 Through specific catalytic mechanisms a Both mechanisms increase the probability that the transition state will actually form LN05 5 c Transition state stabilization i Enzymes active sites are designed in such a way that they make it more favorable for the transition state to form 1 They do this by providing non covalent interactions to the transition state ii Substrates transition states and the active site 1 The active site provides a Substrate specificity b Transition state complementarity i Provides a pull on the substrate leading to the adoption of the electron redistribution in the transition state LN05 6 d Specific catalytic mechanisms i The gist is this 1 Catalytic functional groups on an enzyme may form a transient covalent bond with a compound and activate it for reaction or a group may be transiently transferred from the compound to the enzyme Page 188 ii Three major groups 1 Acid base catalysis a 2 Covalent catalysis a 3 Metal ion catalysis a LN05 7 IV Specific enzymatic reactions pgs 213 216 a The reactions i Enolase 1 This demonstrates Acid base and metal ion catalysis 2 Also transition state stabilization ii Lysozyme 1 This demonstrates covalent catalysis b Enolase i The enzyme and its job Transition state intermediate E S ES ES Base catalysis Metal Ion LN05 8 EP Acid catalysis E P ii Reaction Coordinate Diagram for multiple step process iii iv S G P R C v The reaction and its mechanism 1 General base catalysis and Lys 345 a The Mg cations change the pKa of C 2 making the abstraction possible b Formation of the reaction enolic intermediate c Transition state enolic intermediate stabilization 2 General acid catalysis and Glu 211 Transition state intermediate Reaction intermediate E S ES ES LN05 9 Base catalysis EP Acid catalysis E P c Lysozyme i The enzyme and its job ii The reaction and its mechanism 1 Formation of a covalent bond b w the substrate and enzyme a Covalent catalysis 2 Hydrolysis of this covalent bond regenerating the enzyme Transition Intermediate Reaction State intermediate E S ES ES Acid covalentLN05Base 10catalysis EP E P