X-ray analysis of HIV-I proteinase at 2.7 Å resolution confirms structural homology among retrviral enzymes
Birkbeck, University of London, Londinium, England, United Kingdom Nature
(Impact Factor: 41.46).
12/1989; 342(6247):299-302. DOI: 10.1038/342299a0
Knowledge of the tertiary structure of the proteinase from human immunodeficiency virus HIV-1 is important to the design of inhibitors that might possess antiviral activity and thus be useful in the treatment of AIDS. The conserved Asp-Thr/Ser-Gly sequence in retroviral proteinases suggests that they exist as dimers similar to the ancestor proposed for the pepsins. Although this has been confirmed by X-ray analyses of Rous sarcoma virus and HIV-1 proteinases, these structures have overall folds that are similar to each other only where they are also similar to the pepsins. We now report a further X-ray analysis of a recombinant HIV-1 proteinase at 2.7 A resolution. The polypeptide chain adopts a fold in which the N- and C-terminal strands are organized together in a four-stranded beta-sheet. A helix precedes the single C-terminal strand, as in the Rous sarcoma virus proteinase and also in a synthetic HIV-1 proteinase, in which the cysteines have been replaced by alpha-aminobuytric acid. The structure reported here provides an explanation for the amino acid invariance amongst retroviral proteinases, but differs from that reported earlier in some residues that are candidates for substrate interactions at P3, and in the mode of intramolecular cleavage during processing of the polyprotein.
Available from: Brad Keusch
- "2.3. Molecular dynamics simulations Coordinates for wild-type PR  "
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ABSTRACT: HIV-1 protease (PR) is a 99 amino acid protein responsible for proteolytic processing of the viral polyprotein - an essential step in the HIV-1 life cycle. Drug resistance mutations in PR that are selected during antiretroviral therapy lead to reduced efficacy of protease inhibitors (PI) including darunavir (DRV). To identify the structural mechanisms associated with the DRV resistance mutation L33F, we performed X-ray crystallographic studies with a multi-drug resistant HIV-1 protease isolate that contains the L33F mutation (MDR769 L33F). In contrast to other PR L33F DRV complexes, the structure of MDR769 L33F complexed with DRV reported here displays the protease flaps in an open conformation. The L33F mutation increases noncovalent interactions in the hydrophobic pocket of the PR compared to the wild-type (WT) structure. As a result, L33F appears to act as a molecular anchor, reducing the flexibility of the 30s loop (residues 29-35) and the 80s loop (residues 79-84). Molecular anchoring of the 30s and 80s loops leaves an open S1/S1' subsite and distorts the conserved hydrogen-bonding network of DRV. These findings are consistent with previous reports despite structural differences with regards to flap conformation.
Available from: Jan Konvalinka
- "The major substrate specificity signature features of HIV PR involve (i) a preference for Glu in the P2 0 position; (ii) a certain preference for large aliphatic and Fig. 2. Three-dimensional structures of selected retroviral PRs. (A) Crystal structures of HIV-1 PR in three conformations: the apo-form of the HIV-1 PR with flaps in open conformation (PDB code 3PHV (Lapatto et al., 1989)), HIV-1 PR with flaps in closed conformation (PDB code 4LL3 (Kozisek et al., 2014)) with inhibitor (DRV) bound in the enzyme active site shown in sticks (with carbon atoms colored green, oxygen atoms red, nitrogens blue and sulfur yellow), HIV-1 PR with flaps in semi-open conformation (PDB code 1ZTZ (Cigler et al., 2005). Inhibitor bound in the enzyme active site (metallacarborane) is shown in sticks (with carbon atoms colored yellow, boron atoms cyan, and cobalt atom maroon). "
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ABSTRACT: Proteolytic processing of viral polyproteins is essential for retrovirus infectivity. Retroviral proteases (PR) become activated during or after assembly of the immature, non-infectious virion. They cleave viral polyproteins at specific sites, inducing major structural rearrangements termed maturation. Maturation converts retroviral enzymes into their functional form, transforms the immature shell into a metastable state primed for early replication events, and enhances viral entry competence. Not only cleavage at all PR recognition sites, but also an ordered sequence of cleavages is crucial. Proteolysis is tightly regulated, but the triggering mechanisms and kinetics and pathway of morphological transitions remain enigmatic. Here, we outline PR structures and substrate specificities focusing on HIV PR as a therapeutic target. We discuss design and clinical success of HIV PR inhibitors, as well as resistance development towards these drugs. Finally, we summarize data elucidating the role of proteolysis in maturation and highlight unsolved questions regarding retroviral maturation.
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Available from: Natercia Bras
- "The flexibility and the flap switch in the active site on some aspartic proteases (e.g. HIV-1 protease, renin, BACE1 and plasmepsin II) upon ligands/inhibitors binding were analysed by several experimental and computational studies (Hong & Tang, 2004; Hornak, Okur, Rizzo & Simmerling, 2006; Lapatto et al., 1989; Pietrucci, Marinelli, Carloni, & Laio, 2009; Politi et al., 2011; Sadiq & De Fabritiis, 2010; Steiner et al., 2011; Tzoupis et al., 2012; Xu et al., 2012). Specific MD simulations on HIV-1 protease reveal a reversible transition between open and closed flap conformations (Shang & Simmerling, 2012). "
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ABSTRACT: The aspartic protease renin (REN) catalyses the rate-limiting step in the Renin-Angiotensin-Aldosterone System (RAAS), which regulates cardiovascular and renal homoeostasis in living organisms. Renin blockage is therefore an attractive therapeutic strategy for the treatment of hypertension. Herein, computational approaches were used to provide a structural characterization of the binding site, flap opening and dynamic rearrangements of REN in the key conserved residues and water molecules, with the binding of a dodecapeptide substrate or different inhibitors. All these structural insights during catalysis may assist future studies in developing novel strategies for REN inactivation. Our molecular dynamics simulations of several unbound-REN and bound-REN systems indicate similar flexible-segments plasticity with larger fluctuations in those belonging to the C-domain (exposed to the solvent). These segments are thought to assist the flap opening and closure to allow the binding of the substrate and catalytic water molecules. The unbound-REN simulation suggests that the flap can acquire three different conformations: closed, semi-open and open. Our results indicate that the semi-open conformation is already sufficient and appropriate for the binding of the angiotensinogen (Ang) tail, thus contributing to the high specificity of REN, and that both semi-open and open flap conformations are present in free and complexed enzymes. We additionally observed that the Tyr75-Trp39 H-bond has an important role in assisting flap movement, and we highlight several conserved water molecules and amino acids that are essential for the proper catalytic activity of REN.
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