Shin, JS Yu, MH 2024-01-21T10:42:18Z 2024-01-21T10:42:18Z 2021-09-04 2002-04-05 0021-9258 https://pubs.kist.re.kr/handle/201004/139601 Serpins (serine protease inhibitors) inhibit target proteases by forming a stable covalent complex in which the cleaved reactive site loop of the serpin is inserted into beta-sheet A of the serpin with concomitant translocation of the protease to the opposite of the initial binding site. Despite recent determination of the crystal structures of a Michaelis protease-serpin complex as well as a stable covalent complex, details on the kinetic mechanism remain unsolved mainly due to difficulties in measuring kinetic parameters of acylation, protease translo. cation, and deacylation steps. To address the problem, we applied a mathematical model developed on the basis of a suicide inhibition mechanism to the stopped-flow kinetics of fluorescence resonance energy transfer during complex formation between alpha(1)-antitrypsin, a prototype serpin, and proteases. Compared with the hydrolysis of a peptide substrate, acylation of the protease by alpha(1)-antitrypsin is facilitated, whereas deacylation of the acyl intermediate is strongly suppressed during the protease translocation. The results from nucleophile susceptibility of the acyl intermediate suggest strongly that the active site of the protease is already perturbed during translocation. English AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC PLASMINOGEN-ACTIVATOR INHIBITOR-1 SERPIN-PROTEINASE COMPLEX REACTIVE CENTER LOOP PORCINE PANCREATIC ELASTASE BETA-SHEET-A ALPHA(1)-PROTEINASE INHIBITOR ALPHA-1-PROTEINASE INHIBITOR NATIVE STRAIN CATHEPSIN-G INSERTION Kinetic dissection of alpha(1)-antitrypsin inhibition mechanism Article 10.1074/jbc.M111168200 1 JOURNAL OF BIOLOGICAL CHEMISTRY, v.277, no.14, pp.11629 - 11635 JOURNAL OF BIOLOGICAL CHEMISTRY 277 14 11629 11635 scie scopus 000174846400004 2-s2.0-0037023735 Biochemistry & Molecular Biology Biochemistry & Molecular Biology Article PLASMINOGEN-ACTIVATOR INHIBITOR-1 SERPIN-PROTEINASE COMPLEX REACTIVE CENTER LOOP PORCINE PANCREATIC ELASTASE BETA-SHEET-A ALPHA(1)-PROTEINASE INHIBITOR ALPHA-1-PROTEINASE INHIBITOR NATIVE STRAIN CATHEPSIN-G INSERTION α₁-antitrypsin inhibition mechanism acyl intermediate translocation kinetic model