Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.23.16 (HIV-1 protease)
 2,107 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

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.
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PMID:Observation of a tetrahedral reaction intermediate in the HIV-1 protease-substrate complex. 1579 43

Drug resistance is a very important factor contributing to the failure of current HIV therapies. The ability to understand the resistance mechanism of HIV-protease mutants may be useful in developing more effective and longer lasting treatment regimens. In this paper, we report the first computational study of the clinically relevant E35D mutation of HIV-1 protease in its unbound conformation and complexed with the clinical inhibitor amprenavir and a sample substrate (Thr-Ile-Met-Met-Gln-Arg). Our data, collected from 10 ns molecular-dynamics simulations, show that the E35D mutation results in an increased flexibility of the flaps, thereby affecting the conformational equilibrium between the closed and semi-open conformations of the free protease. The E35D mutation also causes a significant reduction of the calculated binding free energies both for substrate and amprenavir, thus giving a plausible explanation for its ability to increase the level of resistance. One possible explanation for the emergence of this mutation, despite its unfavorable effect on substrate affinity, might be the role of E35D as an escape mutation, which favors escape from the immune system in addition to conferring drug resistance.
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PMID:Insights into amprenavir resistance in E35D HIV-1 protease mutation from molecular dynamics and binding free-energy calculations. 1679 10

As part of our ongoing studies of the human immunodeficiency virus type 1 (HIV-1) protease enzyme, we set out to develop a modular chemical synthesis of the protein from multiple peptide segments. Our initial attempts were frustrated by the insolubility of intermediate peptide products. To overcome this problem, we designed a synthetic strategy combining the solubility-enhancing properties of C-terminal (Arg)n tags and the biological phenomenon of autoprocessing of the Gag-Pol polyprotein that occurs during maturation of the HIV-1 virus in vivo. Synthesis of a 119-residue peptide chain containing 10 residues of the reverse transcriptase (RT) open reading frame plus an (Arg)(10) tag at the C-terminus was straightforward by native chemical ligation followed by conversion of the Cys residues to Ala by Raney nickel desulfurization. The product polypeptide itself completed the final synthetic step by removing the C-terminal modification under folding conditions, to give the mature 99-residue polypeptide. High-purity homodimeric HIV-1 protease protein was obtained in excellent yield and had full enzymatic activity; the structure of the synthetic enzyme was confirmed by X-ray crystallography to a resolution of 1.07 A. This efficient modular synthesis by a biomimetic autoprocessing strategy will enable the facile synthesis of unique chemical analogues of the HIV-1 protease to further elucidate the molecular basis of enzyme catalysis.
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PMID:Modular total chemical synthesis of a human immunodeficiency virus type 1 protease. 1770 84

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 PR monomer and dimer. The D25N mutation (PR(D25N)) increases the equilibrium dimer dissociation constant by a factor >100-fold (1.3 +/- 0.09 microm) 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 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-hydrolyzable substrate analogue, Ac-Thr-Ile-Nle-r-Nle-Gln-Arg-NH(2) (RPB), as compared with PR.DRV and PR.RPB complexes. Although complexation with RPB stabilizes both dimers, the effect on their T(m) is smaller for PR(D25N) (6.2 degrees C) than for PR (8.7 degrees C). The T(m) of PR(D25N).DRV increases by only 3 degrees C relative to free PR(D25N), as compared with a 22 degrees C increase for PR.DRV, and the mutation increases the ligand dissociation constant of PR(D25N).DRV by a factor of approximately 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.
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PMID:Effect of the active site D25N mutation on the structure, stability, and ligand binding of the mature HIV-1 protease. 1828 88

Here we report the X-ray structures of chemically synthesized HIV-1 protease and the inactive [D25N]HIV-1 protease complexed with the ketomethylene isostere inhibitor Ac-Thr-Ile-Nle psi[CO-CH(2)]Nle-Gln-Arg.amide at 1.4 and 1.8A resolution, respectively. In complex with the active enzyme, the keto-group was found to be converted into the hydrated gem-diol, while the structure of the complex with the inactive D25N enzyme revealed an intact keto-group. These data support the general acid-general base mechanism for HIV-1 protease catalysis.
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PMID:Crystal structure of chemically synthesized HIV-1 protease and a ketomethylene isostere inhibitor based on the p2/NC cleavage site. 1899 73

Here we report the X-ray structures of chemically synthesized HIV-1 protease and the inactive [D25N]HIV-1 protease complexed with the ketomethylene isostere inhibitor Ac-Thr-Ile-Nlepsi[CO-CH(2)]Nle-Gln-Arg.amide at 1.4 and 1.8A resolution, respectively. In complex with the active enzyme, the keto-group was found to be converted into the hydrated gem-diol, while the structure of the complex with the inactive D25N enzyme revealed an intact keto-group. These data support the general acid-general base mechanism for HIV-1 protease catalysis.
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PMID:Reprint of "Crystal structure of chemically synthesized HIV-1 protease and a ketomethylene isostere inhibitor based on the p2/NC cleavage site" [Bioorg. Med. Chem. Lett. 18 (2008) 4554-4557]. 1865 69

Structure-based design, synthesis, and biological evaluation of a series of novel HIV-1 protease inhibitors are described. In an effort to enhance interactions with protease backbone atoms, we have incorporated stereochemically defined methyl-2-pyrrolidinone and methyl oxazolidinone as the P1'-ligands. These ligands are designed to interact with Gly-27' carbonyl and Arg-8 side chain in the S1'-subsite of the HIV protease. We have investigated the potential of these ligands in combination with our previously developed bis-tetrahydrofuran (bis-THF) and cyclopentanyltetrahydrofuran (Cp-THF) as the P2-ligands. Inhibitor 19b with a (R)-aminomethyl-2-pyrrolidinone and a Cp-THF was shown to be the most potent compound. This inhibitor maintained near full potency against multi-PI-resistant clinical HIV-1 variants. A high resolution protein-ligand X-ray crystal structure of 19b-bound HIV-1 protease revealed that the P1'-pyrrolidinone heterocycle and the P2-Cp-ligand are involved in several critical interactions with the backbone atoms in the S1' and S2 subsites of HIV-1 protease.
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PMID:Design of HIV-1 protease inhibitors with pyrrolidinones and oxazolidinones as novel P1'-ligands to enhance backbone-binding interactions with protease: synthesis, biological evaluation, and protein-ligand X-ray studies. 1947 17

The binding mechanism of a peptide substrate (Thr-Ile-Met-Met-Gln-Arg, cleavage site p2-NC of the viral polyprotein) to wild-type HIV-1 protease has been investigated by 1.6 micros biased all-atom molecular dynamics simulations in explicit water. The configuration space has been explored biasing seven reaction coordinates by the bias-exchange metadynamics technique. The structure of the Michaelis complex is obtained starting from the substrate outside the enzyme within a backbone rmsd of 0.9 A. The calculated free energy of binding is -6 kcal/mol, and the kinetic constants for association and dissociation are 1.3 x 10(6) M(-1) s(-1) and 57 s(-1), respectively, consistent with experiments. In the main binding pathway, the flaps of the protease do not open sizably. The substrate slides inside the enzyme cavity from the tight lateral channel. This may contrast with the natural polyprotein substrate which is expected to bind by opening the flaps. Thus, mutations might influence differently the binding kinetics of peptidomimetic ligands and of the natural substrate.
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PMID:Substrate binding mechanism of HIV-1 protease from explicit-solvent atomistic simulations. 1964 90

Plasmodium falciparum plasmepsin V (PfPMV) is an essential aspartic protease required for parasite survival, thus, considered as a potential drug target. This study reports the first detailed structural analysis and molecular dynamics simulation of PfPMV as an apoenzyme and its complexes with the substrate PEXEL as well as with the inhibitor saquinavir. The presence of pro-peptide in PfPMV may not structurally hinder the formation of a functionally competent catalytic active site. The structure of PfPMV-PEXEL complex shows that the unique positions of Glu179 and Gln222 are responsible for providing the specificity of PEXEL substrate with arginine at P3 position. The structural analysis also reveals that the S4 binding pocket in PfPMV is occupied by Ile94, Ala98, Phe370 and Tyr472, and therefore, does not allow binding of pepstatin, a potent inhibitor of most pepsin-like aspartic proteases. Among the screened inhibitors, the HIV-1 protease inhibitors and KNI compounds have higher binding affinities for PfPMV with saquinavir having the highest value. The presence of a flexible group at P2 and a bulky hydrophobic group at P3 position of the inhibitor is preferred in the PfPMV substrate binding pocket. Results from the present study will aid in the design of potent inhibitors of PMV.
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PMID:Understanding the structural basis of substrate recognition by Plasmodium falciparum plasmepsin V to aid in the design of potent inhibitors. 2753 85

Multidrug resistance to current Food and Drug Administration-approved HIV-1 protease (PR) inhibitors drives the need to understand the fundamental mechanisms of how drug pressure-selected mutations, which are oftentimes natural polymorphisms, elicit their effect on enzyme function and resistance. Here, the impacts of the hinge-region natural polymorphism at residue 35, glutamate to aspartate (E35D), alone and in conjunction with residue 57, arginine to lysine (R57K), are characterized with the goal of understanding how altered salt bridge interactions between the hinge and flap regions are associated with changes in structure, motional dynamics, conformational sampling, kinetic parameters, and inhibitor affinity. The combined results reveal that the single E35D substitution leads to diminished salt bridge interactions between residues 35 and 57 and gives rise to the stabilization of open-like conformational states with overall increased backbone dynamics. In HIV-1 PR constructs where sites 35 and 57 are both mutated (e.g. E35D and R57K), x-ray structures reveal an altered network of interactions that replace the salt bridge thus stabilizing the structural integrity between the flap and hinge regions. Despite the altered conformational sampling and dynamics when the salt bridge is disrupted, enzyme kinetic parameters and inhibition constants are similar to those obtained for subtype B PR. Results demonstrate that these hinge-region natural polymorphisms, which may arise as drug pressure secondary mutations, alter protein dynamics and the conformational landscape, which are important thermodynamic parameters to consider for development of inhibitors that target for non-subtype B PR.
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PMID:Effects of Hinge-region Natural Polymorphisms on Human Immunodeficiency Virus-Type 1 Protease Structure, Dynamics, and Drug Pressure Evolution. 2757 89


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