3bvb

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Template:STRUCTURE 3bvb

Cystal structure of HIV-1 Active Site Mutant D25N and inhibitor Darunavir

Overview

All aspartic proteases including retroviral proteases share the triplet DTG critical for the active site geometry and catalytic function. These residues interact closely in the active, dimeric structure of HIV-1 protease (PR). We have systematically assessed the effect of the D25N mutation on the structure and stability of the mature HIV-1 protease (PR) monomer and dimer. The D25N mutation (PR(D25N)) increases the equilibrium dimer dissociation constant by a factor >100-fold (1.3 +/- 0.09 muM) relative to PR. In the absence of inhibitor, NMR studies reveal clear structural differences between PR and PR(D25N) in the relatively mobile P1 loop (residues 79-83) and flap regions, and differential scanning calorimetric (DSC) analyses show that the mutation lowers the stabilities of both the monomer and dimer folds by 5 and 7.3 degrees C, respectively. Only minimal differences are observed in high resolution crystal structures of PR(D25N) complexed to darunavir (DRV), a potent clinical inhibitor, or a non-hydrolysable substrate analogue (RPB), as compared to PR-DRV and PR-RPB complexes. Although complexation with RPB stabilizes both dimers, the effect on their Tm is smaller for PR(D25N) (6.2 degrees C) than for PR (8.7 degrees C). The Tm of PR(D25N)/DRV increases by only 3 degrees C relative to free PR(D25N), as compared to a 22 degrees C increase for PR/DRV, and the mutation increases the ligand dissociation constant of PR(D25N)/DRV by a factor of ~10(6) relative to PR/DRV. These results suggest that interactions mediated by the catalytic Asp residues make a major contribution to the tight binding of DRV to PR.

About this Structure

3BVB is a Single protein structure of sequence from Human immunodeficiency virus 1. Full crystallographic information is available from OCA.

Reference

Effect of the active-site D25N mutation on the structure, stability and ligand binding of the mature HIV-1 protease., Sayer JM, Liu F, Ishima R, Weber IT, Louis JM, J Biol Chem. 2008 Feb 15;. PMID:18281688 Page seeded by OCA on Sun May 4 21:08:59 2008