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B0. Introduction to Enzymatic Mechanisms

  • Page ID
    9062
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    We can apply what we learned about catalysis by small molecules to enzyme-catalyzed reactions. To understand the mechanism of an enzyme-catalyzed reaction, we try to alter as many variables, one at a time, and ascertain the effects of the changes on the activity of the enzyme. Kinetic methods can be used to obtain data from which inferences about the mechanism can be made. Obviously, crystal structures of the enzyme in the presence and absence of a competitive inhibitor give abundant information about possible mechanisms. It is amazing, however, how much information about enzyme mechanism can be gained even if all you have is a blender, a stopwatch, an impure enzyme, and a few substrates and inhibiting reagents. Systematically, the kineticist, medicinal chemist and molecular biologists (i.e. a well trained chemist) can change:

    1. the substrate - for example, changing the leaving group or acyl sustituents of a hydrolyzable substrate;
    2. the pH or ionic strength - which can give data about general acids/bases in the active site;
    3. the enzyme - by chemical modification of specific amino acids, or through site-specific mutagenesis;
    4. the solvent - as will be discussed in the next chapter section .

    We will explore in detail the mechanisms of three enzymes. For carboxypeptidase, we will study possible mechanisms for the cleavage of C-terminal hydrophobic amino acids from a peptide. For lysozyme, we will study the structural features of the enzyme and substrates along with the mechanism for cleavage of glycosidic links in bacterial peptidoglycan cell walls. For chymotrypsin, we will study experiments which vary the substrate, pH, and the enzyme and infer from this information about a mechanism consistent with the experimental data. Kinetic analyses can be used to determine the:

    • order of binding/dissociation of substrates and products
    • rate constants for individual steps
    • and clues to the nature of catalytic groups found in the enzyme.

    This page titled B0. Introduction to Enzymatic Mechanisms is shared under a CC BY-NC-SA license and was authored, remixed, and/or curated by Henry Jakubowski.

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