Abstract
In the preceding study, mutant Rous sarcoma virus (RSV) proteases are described in which three amino acids found in the human immunodeficiency virus-1 (HIV-1) protease (PR) were substituted into structurally comparable positions (Grinde, B., Cameron, C.E., Leis, J., Weber, I., Wlodawer, A., Burstein, H., Bizub, D., and Skalka, A. M. (1992) J. Biol. Chem. 267, 9481-9490). In this report, the activity of the wild type and these mutant PRs are compared using a set of RSV NC-PR peptide substrates with single amino acid substitutions in each of the P4 to P3' positions. With most substrates, the relative activities of the two active mutants followed that of the RSV PR. Substitutions in the P1 and P1' positions were an exception; in this case, the mutants behaved more like the HIV-1 PR. These results confirm predictions from structural analyses which indicate that residues 105 and 106 of the RSV PR are important in forming the S1 and S1' binding subsites. These results, further analyzed with the aid of computer modeling of the RSV PR with different substrates, provide an explanation for why only partial HIV-1 PR-like behavior was introduced into the above RSV PR mutants.
Highlights
Haim Burstein and Anna MariSekalka From the Fox Chase Cancer Center, Institutefor Cancer Research, Philadelphia, Pennsylvania19111
Mutant Rous sarcomavirus currently available. These include the native structure of the (RSV) proteases are described in which three amino RSV PR, refined at 2-A resolution ( l ), and three separate acids found in the human immunodeficiency virus-1 (HIV-1)protease (PR) were substituted into structurally comparable positions
Molecular Modeling of the Flaps of the RSV PR-Several residues, including N61 to G69, from one subunit and A59’ to G69’ from the other subunit in each flap of the RSV PR dimer are not visible in the crystal structure and are considered to be disordered. (Note that the prime notation in this context is used to denote the second subunit of the P R dimer.) Since the flap region is presumed to be very important for substrate binding and catalysis, and crystal structures of the RSV PR complexed to ligand arenot available, we have modeled the structure of the RSV PR flaps based on the cocrystal structure of HIV-1 PR with the inhibitor MVTlOl [6]
Summary
Amino acids and numbers written in efficiencies by both the AMV and HIV-1 wild type PRs as well as the mutant RSV PRs [13]. The activity of the two mutantand twowild type PRs were examined using 40 different peptide substrates with single amino acid changes placed in the P4through the P3’ positions. Results from this analysis show that amino acid substitutions in the PR at positions 105 and 106, whichare primarily part of the S1and S1‘ enzyme subsites, cause changes which affect almost exitalics refer to theHIV-1 PR. Preparationof HIV-1 PR, from E. coli, and avian myeloblastosis virus (AMV) PR, from virions, has been described [16, 17]. Molecular Modeling-Atomic coordinates of RSV PR [1, 18] and HIV-1 PR with inhibitor [6] were examined on an Evans and Sutherland PS390 computer graphics system using the program FRODO [19]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
