You are a biochemist researching the mechanism of chymotrypsin catalyzed peptide
ID: 3478574 • Letter: Y
Question
You are a biochemist researching the mechanism of chymotrypsin catalyzed peptide bond cleavage. You engineer a chymotrypsin mutant that substitutes an Arginine for the active site Histidine (residue #57) normally found in chymotrypsin. Specifically describe the effect that this mutation would have on the molecular mechanism of the reaction as well as the effect it would have on the kcat and Km of chymotrypsin (Will the Arginine substituted enzyme result in proper catalysis or not? If yes, what are the mechanistic differences exhibited by the mutant form of chymotrypsin. If no, why not and at what point would you expect the mechanism to "fail"). My hint to you is that Arginine is a great specific base, but a terrible specific acid. (Limit your response to 5 sentences or less).Explanation / Answer
Arginine PI is 10.76 that is quite more basic than the histidine has 7.59. The arginine substitution greatly increases the basic nature of the catalytic triad that made of His-57, Asp-102, and Ser-195. In chymotrypsin, the histidine work for proton transfer, after arginine substitution, the catalytic domain has to increase their more protonation pH for releasing the proton.
Therefore, the protein after get substitution cannot work. The Kcat will drastically decrease and Km will too drastically increase.
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