Human immunodeficiency virus 1 (HIV-1) protease is an aspartyl protease composed of two identical protomers linked by a four-stranded antiparallel β-sheet consisting of the NH2- and COOH-terminal segments (Weber, I. T. (1990) J. Biol. Chem. 265, 10492-10496). Kinetic analysis of the HIV-1 protease-catalyzed hydrolysis of a fluorogenic substrate demonstrates that the enzyme is an obligatory dimer. At pH = 5.0, 0.1 M sodium acetate, 1 M NaCl, 1 mM EDTA buffer, 37°C, the equilibrium dissociation constant, K(d) = 3.6 ± 1.9 nM. We found that the tetrapeptide Ac-Thr-Leu-Asn-Phe-COOH, corresponding to the COOH-terminal segment of the enzyme, is an excellent inhibitor of the enzyme. Kinetic analysis shows that the inhibitor binds to the inactive protomers and prevents their association into the active dimer (dissociative inhibition). The dissociative nature of this inhibition is consistent with the results obtained from sedimentation equilibrium experiments in which the apparent molecular weight of the enzyme was observed to be 20,800 ± 1,500 and 12,100 ± 300, in the absence and presence of the COOH-terminal tetrapeptide, respectively. The dissociation constant of the protomer-inhibitor complex is K(i) = 45.1 ± 1.8 μM. This is the first kinetic analysis and direct experimental demonstration of noncovalent dissociative inhibition.
|Original language||English (US)|
|Number of pages||4|
|Journal||Journal of Biological Chemistry|
|State||Published - Oct 29 1991|
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology