title: Design of protease inhibitors on the basis of substrate stereospecificity.
abstract: The substrate stereospecificity in enzymic reactions, which is one of characteristics of enzymes along with the substrate and regiospecificity can provide a basis for the rational design of inhibitors. This has been demonstrated using alpha-chymotrypsin, a prototypic serine protease as a model enzyme. On the basis of the structure-activity relationships for substrates as well as inhibitors and mechanism of the enzymic reaction, a schematic three-dimensional model of the S1 subsite of alpha-chymotrypsin is constructed. It was envisioned from the three-dimensional active site model that 2-benzyl-3,4-epoxybutanoic acid methyl ester (1) having a (2S)-configuration would bind the enzyme with its oxirane ring being rested at the catalytic site, in which the oxirane ring is subject to a nucleophilic attack by the Ser-195 hydroxyl to form a ether linkage. Kinetic analysis of the enzymic reaction in the presence of the potential inhibitors showed that (2S,3R)-1 inactivates alpha-chymotrypsin, while (2S,3S)-1 inhibits the enzyme competitively. The lack of inactivating activity in the case of (2S,3S)-1 may be due to the unfavorable alignment of the C3-O bond with respect to the hydroxyl of Ser-195 for the SN2-type ring cleave reaction of the oxirane moiety. When the design protocol was applied to papain, a representative cysteine protease, (2S,3S)-1 inhibited the enzyme irreversibly, while (2S,3R)-1 inhibited reversibly. On the basis of the stereospecificity shown in the inactivation of the enzymes, it was inferred that in the case of alpha-chymotrypsin, the nucleophilic attack of the Ser-195 hydroxyl at the scissile carbonyl carbon of substrates occurs in a si fashion, while the thiolate of Cys-25 in papain attacks the substrate amide bond in a re fashion. The inhibitor design protocol may be applied to other proteases.
authors.affiliation: Center for Biofunctional Molecules, Pohang University of Science and Technology, Korea.
authors.forename: D H
authors.initials: DH
authors.lastname: Kim
mesh_terms: D001665:Binding Sites; D002918:Chymotrypsin / Q000737:chemistry* / Q000378:metabolism; D015195:Drug Design; D010206:Papain / Q000037:antagonists & inhibitors* / Q000737:chemistry; D011480:Protease Inhibitors / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology; D013237:Stereoisomerism; D013379:Substrate Specificity
chemical_list: D011480:Protease Inhibitors; D002918:Chymotrypsin; D010206:Papain