ACTIVE-SITE MOBILITY IN HUMAN-IMMUNODEFICIENCY-VIRUS, TYPE-1, PROTEASE AS DEMONSTRATED BY CRYSTAL-STRUCTURE OF A28S MUTANT

The mutation Ala(28) to serine in human immunodeficiency virus, type 1 , (HIV-1) protease introduces putative hydrogen bonds to each active-s ite carboxyl group. These hydrogen bonds are ubiquitous in pepsin-like eukaryotic aspartic proteases. In order to understand the significanc e of this difference between HIV-1 protease and homologous, eukaryotic aspartic proteases, we solved the three-dimensional structure of A28S mutant HIV-1 protease in complex with a peptidic inhibitor U-89360E. The structure has been determined to 2.0 Angstrom resolution with an R factor of 0.194. Comparison of the mutant enzyme structure with that of the wild-type HN-I protease bound to the same inhibitor (Hong L, Tr eharne A, Hartsuck JA, Foundling S, Tang J, 1996, Biochemistry 35:1062 7-10633) revealed double occupancy for the Ser(28) hydroxyl group, whi ch forms a hydrogen bond either to one of the oxygen atoms of the acti ve-site carboxyl or to the carbonyl oxygen of Asp(30). We also observe d marked changes in orientation of the Asp(25) catalytic carboxyl grou ps, presumably caused by the new hydrogen bonds. These observations su ggest that catalytic aspartyl groups of HIV-1 protease have significan t conformational flexibility unseen in eukaryotic aspartic proteases. This difference may provide an explanation for some unique catalytic p roperties of HIV-1 protease.