Effect of Nepsilon-alkylation of the substrate on the hydrolysis of benzoylglycyl-L-lysine by carboxypeptidase B1.
The action of carboxypeptidase B (EC 3.4.12.3) on the benzoylglycyl dipeptides Bz-Gly-Lys(X) where X = methyl, ethyl, propyl, formyl, dimethyl, isopropyl, trimethyl, and benzyl has been investigated. All were hydrolyzed, at a rate decreasing in the order indicated, except where X = trimethyl and benzyl. Compounds where X = dimethyl, formyl, and isopropyl were hydrolyzed very slowly, and did not inhibit the hydrolysis of Bz-Glys-Lys by the enzyme. The kinetic parameters kcat and Km were determined for compounds with X = methyl and ethyl. The observed decrease in rate of hydrolysis of substrates with increasing size of X is consistent with increasing steric hindrance effects arising from the interaction of the Nepsilon-alkyl group with residues of the protein in the cleft which accommodates the substrate side-chain, and resulting in weaker binding of the substrate. Bz-Gly-Lys(Me2) was prepared by reductive methylation of Bz-Gly-Lys. Bz-Gly-Lys(Me3) was prepared by the reaction of Bz-Gly-Lys(Me2) with diazomethane in aqueous solution. Bz-Gly-Lys(Me) and Bz-Gly-Lys(Et) were synthesized by classical coupling procedures from the appropriately protected lysine derivatives.
D000438:Alcohols; D000478:Alkylation; D000818:Animals; D001565:Benzoates; D002268:Carboxypeptidases / Q000378:metabolism*; D004151:Dipeptides; D005998:Glycine; D007700:Kinetics; D008239:Lysine; D013329:Structure-Activity Relationship; D013552:Swine
D000438:Alcohols; D001565:Benzoates; D004151:Dipeptides; D002268:Carboxypeptidases; D008239:Lysine; D005998:Glycine