title: Observation of a tetrahedral reaction intermediate in the HIV-1 protease-substrate complex.
abstract: HIV-1 protease is an effective target for the design of drugs against AIDS. To help this process of drug design, three-dimensional structures have been determined of complexes between HIV-1 protease and a variety of transition-state analogue inhibitors. The true transition state, however, has not been structurally characterized. The crystal structure of the C95M/C1095A HIV-1 protease tethered dimer shows a distinctive feature in which the two flaps of the enzyme are in a 'closed conformation' even in the unliganded state. This unique feature has been utilized here to study the structure of HIV-1 protease complexed to an oligopeptide substrate of amino acid sequence His-Lys-Ala-Arg-Val-Leu*NPhe-Glu-Ala-Nle-Ser (where * denotes the cleavage site, and NPhe and Nle denote p-nitrophenylalanine and norleucine residues respectively). The X-ray structure of the complex refined against 2.03 A (0.203 nm) resolution synchrotron data shows that the substrate is trapped as a tetrahedral reaction intermediate in the crystal. The hydrogen-bonding interactions between the reaction intermediate and the catalytic aspartates are different from those observed previously using transition-state analogues. The reaction intermediate did not dissociate to release the products, possibly due to the inflexibility introduced in the flaps when the enzyme is packed inside crystals.
authors.affiliation: Solid State Physics Division, Bhabha Atomic Research Centre, Trombay, Mumbai-400085, India.
authors.forename: Mukesh
authors.initials: M
authors.lastname: Kumar
authors.forename: Vishal
authors.initials: V
authors.lastname: Prashar
authors.forename: Smita
authors.initials: S
authors.lastname: Mahale
authors.forename: Madhusoodan V
authors.initials: MV
authors.lastname: Hosur
mesh_terms: D001665:Binding Sites; D018360:Crystallography, X-Ray; D016333:HIV Protease / Q000737:chemistry* / Q000378:metabolism*; D006860:Hydrogen Bonding; D008958:Models, Molecular; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D011487:Protein Conformation; D055672:Static Electricity; D013379:Substrate Specificity
chemical_list: D010455:Peptides; D016333:HIV Protease