3BXR

Crystal Structures Of Highly Constrained Substrate And Hydrolysis Products Bound To HIV-1 Protease. Implications For Catalytic Mechanism

Summary for 3BXR

• Entry DOI: 10.2210/pdb3bxr/pdb
• Related: 1B6P
• Descriptor: Protease, SULFATE ION, (9S,12S)-9-(1-methylethyl)-N-[(8S,11S)-8-[(1S)-1-methylpropyl]-7,10-dioxo-2-oxa-6,9-diazabicyclo[11.2.2]heptadeca-1(15),13,16-trien-11-yl]-7,10-dioxo-2-oxa-8,11-diazabicyclo[12.2.2]octadeca-1(16),14,17-triene-12-carboxamide, ... (4 entities in total)
• Functional Keywords: hiv protease, hivpr, substrate, product, aids, aspartyl protease, capsid maturation, core protein, cytoplasm, dna integration, dna recombination, dna-directed dna polymerase, endonuclease, hydrolase, lipoprotein, magnesium, membrane, metal-binding, multifunctional enzyme, myristate, nuclease, nucleotidyltransferase, nucleus, phosphoprotein, rna-binding, rna-directed dna polymerase, transferase, viral nucleoprotein, virion, zinc, zinc-finger
• Cellular location: Gag-Pol polyprotein: Host cell membrane; Lipid-anchor. Matrix protein p17: Virion membrane; Lipid- anchor . Capsid protein p24: Virion . Nucleocapsid protein p7: Virion . Reverse transcriptase/ribonuclease H: Virion . Integrase: Virion : P03369
• Total number of polymer chains: 2
• Total formula weight: 22591.48

Authors

Tyndall, J.D.,Pattenden, L.K.,Reid, R.C.,Hu, S.H.,Alewood, D.,Alewood, P.F.,Walsh, T.,Fairlie, D.P.,Martin, J.L. (deposition date: 2008-01-14, release date: 2008-03-25, Last modification date: 2023-11-15)

Primary citation

Tyndall, J.D.,Pattenden, L.K.,Reid, R.C.,Hu, S.H.,Alewood, D.,Alewood, P.F.,Walsh, T.,Fairlie, D.P.,Martin, J.L.
Crystal Structures of Highly Constrained Substrate and Hydrolysis Products Bound to HIV-1 Protease. Implications for the Catalytic Mechanism
Biochemistry, 47:3736-3744, 2008

PubMed Abstract: HIV-1 protease is a key target in treating HIV infection and AIDS, with 10 inhibitors used clinically. Here we used an unusual hexapeptide substrate, containing two macrocyclic tripeptides constrained to mimic a beta strand conformation, linked by a scissile peptide bond, to probe the structural mechanism of proteolysis. The substrate has been cocrystallized with catalytically active synthetic HIV-1 protease and an inactive isosteric (D25N) mutant, and three-dimensional structures were determined (1.60 A). The structure of the inactive HIVPR(D25N)/substrate complex shows an intact substrate molecule in a single orientation that perfectly mimics the binding of conventional peptide ligands of HIVPR. The structure of the active HIVPR/product complex shows two monocyclic hydrolysis products trapped in the active site, revealing two molecules of the N-terminal monocyclic product bound adjacent to one another, one molecule occupying the nonprime site, as expected, and the other monocycle binding in the prime site in the reverse orientation. The results suggest that both hydrolysis products are released from the active site upon cleavage and then rebind to the enzyme. These structures reveal that N-terminal binding of ligands is preferred, that the C-terminal site is more flexible, and that HIVPR can recognize substrate shape rather than just sequence alone. The product complex reveals three carboxylic acids in an almost planar orientation, indicating an unusual hexagonal homodromic complex between three carboxylic acids. The data presented herein regarding orientation of catalytic aspartates support the cleavage mechanism proposed by Northrop. The results imply strategies for design of inhibitors targeting the N-terminal side of the cleavage site or taking advantage of the flexibility in the protease domain that accommodates substrate/inhibitor segments C-terminal to the cleavage site.

PubMed: 18311928
DOI: 10.1021/bi7023157

Experimental method

X-RAY DIFFRACTION (1.6 Å)