MCB 450 1st Edition Lecture 12 Outline of Last Lecture I Effect of substrate and enzyme on reaction rate II Km and Vmax III Kcat IV kcat Km V Bi substrate reactions Outline of Current Lecture I Catalytic strategies used by enzymes II Inhibition of enzyme activity III Example of an enzyme mechanism a peptide hydrolysis by chymotrypsin Current Lecture Effect of heat on enzyme activity As you heat up the enzyme it will eventually reach a point where it will be too hot and will denature Effect of pH on reaction rate As the pH increases we start to see a bell shaped curve because the enzyme will become deprotonated Enzyme won t work if amino acid 1 or 2 is deprotonated There is a specific active state that specifies if an amino acid must be protonated or deprotonated to work Inhibition of enzyme activity When we talk about enzymes inhibitors we are talking about very small molecules Competitive inhibition Shows formulation for enzyme and substrate along with the formulation for the enzyme and inhibitor o When the inhibitor reacts with the enzyme it creates an enzyme inhibitor complex Effect of a competitive inhibitor on Km and Vmax Getting less and less of a competitive inhibitor o Actually overcoming the competitive inhibitor as the reaction goes on with a high substrate reaction When using a competitive inhibitor Vmax will stay the same and Km will go up These notes represent a detailed interpretation of the professor s lecture GradeBuddy is best used as a supplement to your own notes not as a substitute Uncompetitive inhibition Actually lowering the Km for the enzyme Vmax also decreases With competitive inhibitor the intercepts are the same but with uncompetitive inhibitor the intercepts are different Noncompetitive inhibition Noncompetitive inhibition causes the Vmax to decreases but the Km remains the same Solving problems on enzyme inhibition Could be something on the upcoming exam Will have to calculate apparent Km in the homework Know the three types of reversible inhibitors effects on Km and Vmax Acetylcholinesterase Mechanism of this enzyme is very similar to chymotrypsin Chemical messenger in your nerve transmission o Acetylcholinesterase 2 phases of the chymotrypsin reaction Chymotrypsin reaction has two steps o Ser 195 and oxygen from water will both act as nucleophiles If you keep track of the His 57 you can keep track of what is happening in the chymotrypsin reaction Catalytic triad Shows how catalytic triad works Oxyanion hole Shows negatively charged oxygen on the carbonyl Interactions in oxyanion hole also contributes to the binding energy that stabilize the transition state formed on the way to the tetrahedral intermediate H bonds between negatively O and peptide NH groups in the peptide backbone Chymotrypsin mechanism dacylation Reversal of acylation of the serine in the enzyme o Deacylation Histidine can now act as a general acid His 57 functions as general base acid Summary of the roles of histidine in acylation and deacylation Serine proteases example of a ping pong reaction Now can we get evidence for formation of an acyl enzyme intermediate Getting evidence for formation of an acyl enzyme intermediate in the chymotrypsin reaction Basically just want to slow the reaction down and then see if you can detect the two steps