MCB 450 1st Edition Lecture 12Outline of Last Lecture I. Effect of [substrate] and [enzyme] on reaction rateII. Km and VmaxIII. KcatIV. kcat / KmV. Bi-substrate reactionsOutline of Current Lecture I. Catalytic strategies used by enzymesII. Inhibition of enzyme activityIII. Example of an enzyme mechanism:a. peptide hydrolysis by chymotrypsinCurrent LectureEffect 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 denatureEffect of pH on reaction rate-As the pH increases we start to see a bell-shaped curve because the enzyme will becomedeprotonated-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 moleculesCompetitive inhibition-Shows formulation for enzyme and substrate along with the formulation for the enzyme and inhibitoroWhen the inhibitor reacts with the enzyme it creates an enzyme-inhibitor complexEffect of a competitive inhibitor on Km and Vmax-Getting less and less of a competitive inhibitoroActually overcoming the competitive inhibitor as the reaction goes on with a highsubstrate reaction-When using a competitive inhibitor Vmax will stay the same and Km will go upThese 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 differentNoncompetitive inhibition-Noncompetitive inhibition causes the Vmax to decreases but the Km remains the sameSolving 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 VmaxAcetylcholinesterase-Mechanism of this enzyme is very similar to chymotrypsin-Chemical messenger in your nerve transmissionoAcetylcholinesterase2 phases of the chymotrypsin reaction-Chymotrypsin reaction has two stepsoSer-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 reactionCatalytic triad-Shows how catalytic triad worksOxyanion 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 backboneChymotrypsin mechanism: dacylation-Reversal of acylation of the serine in the enzymeoDeacylation-Histidine can now act as a general acidHis-57 functions as general base & acid-Summary of the roles of histidine in acylation and deacylationSerine 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