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)

Site-directed mutagenesis of autolysis sites in the human immunodeficiency virus type 1 (HIV-1) protease was applied in an analysis of enzyme specificity; the protease served, therefore, as both enzyme and substrate in this study. Inspection of natural substrates of all retroviral proteases revealed the absence of beta-branched amino acids at the P1 site and of Lys anywhere from P2 through P2'. Accordingly, several mutants of the HIV-1 protease were engineered in which these excluded amino acids were substituted at their respective P positions at the three major sites of autolysis in the wild-type protease (Leu5-Trp6, Leu33-Glu34, and Leu63-Ile64), and the mutant enzymes were evaluated in terms of their resistance to autodegradation. All of the mutant HIV-1 proteases, expressed as inclusion bodies in Escherichia coli, were enzymatically active after refolding, and all showed greatly diminished rates of cleavage at the altered autolysis sites. Some, however, were not viable enzymatically because of poor physical characteristics. This was the case for mutants having Lys replacements of Glu residues at P2' and for another in which all three P1 leucines were replaced by Ile. However, one of the mutant proteases, Q7K/L33I/L63I, was highly resistant to autolysis, while retaining the physical properties, specificity, and susceptibility to inhibition of the wild-type enzyme. Q7K/L33I/L63I should find useful application as a stable surrogate of the HIV-1 protease. Overall, our results can be interpreted relative to a model in which the active HIV-1 protease dimer is in equilibrium with monomeric, disordered species which serve as the substrates for autolysis.
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PMID:The HIV-1 protease as enzyme and substrate: mutagenesis of autolysis sites and generation of a stable mutant with retained kinetic properties. 806 16

A vector projection method is proposed to predict the cleavability of oligopeptides by extended-specificity site proteases. For an enzyme with eight specificity subsites the substrate octapeptide can be uniquely expressed as a vector in an 8-dimensional space, whose eight bases correspond to the amino acids at the eight subsites, P4, P3, P2, P1, P1', P2', P3', and P4', respectively. The component of such a characteristic vector on each of the eight bases is defined as the frequency of an amino acid occurring at a given site. These frequencies were derived from a set of octapeptides known to be cleaved by HIV protease. The cleavability of an octapeptide can then be estimated from the projection of its characteristic vector on an idealized, optimally cleavable vector. The high ratio of correct prediction vs. total prediction for the data in both the training and the testing sets indicates that the new method is self-consistent and efficient. It provides a rapid and accurate algorithm for analyzing the specificity of any multi-subsite enzyme for which there is no coupling between subsites. In particular, it is useful for predicting the cleavability of an oligopeptide by either HIV-1 or HIV-2 protease, and hence offers a supplementary means for finding effective inhibitors of HIV protease as potential drugs against AIDS.
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PMID:A vector projection approach to predicting HIV protease cleavage sites in proteins. 833 7

C7 mimetics, designed to lock three amino acid residues of a peptide chain into a gamma-turn conformation, were introduced sequentially between the P3 to P2' positions of a model HIV-1 protease substrate I (resulting in compounds II-IV) to probe its conformational requirements in binding to HIV-1 protease. Of these, compound IIIa with the C7 mimetic replacing Asn-Tyr-Pro, corresponding to the P2 through P1' positions of substrate, was found to be an inhibitor with a Ki of 147 microM. Reduction of the amide bond in the C7 mimetic of IIIa resulted in a novel constrained reduced-amide mimetic VIa with a Ki of 430 nM. This corresponds to over a 300-fold improvement in inhibitory activity over the original C7 mimetic. The inhibitory activity of mimetic VIa was in addition found to be 44-fold better than a similar linear reduced-amide containing inhibitor V. The synthesis of these mimetics are described.
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PMID:A novel constrained reduced-amide inhibitor of HIV-1 protease derived from the sequential incorporation of gamma-turn mimetics into a model substrate. 836 Aug 76

The design, synthesis, and molecular modeling studies of a novel series of azacyclic ureas, which are inhibitors of human immunodeficiency virus type 1 (HIV-1) protease that incorporate different ligands for the S1', S2, and S2' substrate-binding sites of HIV-1 protease are described. The synthesis of this series is highly flexible in the sense that the P1', P2, and P2' residues of the inhibitors can be changed independently. Molecular modeling studies on the phenyl ring of the P2 and P2' ligand suggested incorporation of hydrogen-bonding donor/acceptor groups at the 3' and 4-positions of the phenyl ring should increase binding potency. This led to the discovery of compound 7f (A-98881), which possesses high potency in the HIV-1 protease inhibition assay and the in vitro MT-4 cell culture assay (Ki = approximately 5 pM and EC50 = 0.002 microM). This compares well with the symmetrical cyclic urea 1 pioneered at DuPont Merck.
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PMID:A novel, picomolar inhibitor of human immunodeficiency virus type 1 protease. 855 7

A series of novel, azacyclic ureas which are highly potent inhibitors of the HIV-1 protease (IC50 = 4.1 to < 0.5 nM) were synthesized. Aqueous solubilities of this series of compounds were improved by incorporating polar functional groups at the P1' P2 and P2' positions. These compounds also possess good anti-viral activity by inhibition of the cytopathic effect of HIV-13B in MT-4 cells in vitro.
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PMID:Novel azacyclic ureas that are potent inhibitors of HIV-1 protease. 875 80

High-resolution X-ray structures of the complexes of HIV-1 protease (HIV-1PR) with peptidomimetic inhibitors reveal the presence of a structural water molecule which is hydrogen bonded to both the mobile flaps of the enzyme and the two carbonyls flanking the transition-state mimic of the inhibitors. Using the structure-activity relationships of C2-symmetric diol inhibitors, computed-aided drug design tools, and first principles, we designed and synthesized a novel class of cyclic ureas that incorporates this structural water and preorganizes the side chain residues into optimum binding conformations. Conformational analysis suggested a preference for pseudodiaxial benzylic and pseudodiequatorial hydroxyl substituents and an enantiomeric preference for the RSSR stereochemistry. The X-ray and solution NMR structure of the complex of HIV-1PR and one such cyclic urea, DMP323, confirmed the displacement of the structural water. Additionally, the bound and "unbound" (small-molecule X-ray) ligands have similar conformations. The high degree of preorganization, the complementarity, and the entropic gain of water displacement are proposed to explain the high affinity of these small molecules for the enzyme. The small size probably contributes to the observed good oral bioavailability in animals. Extensive structure-based optimization of the side chains that fill the S2 and S2' pockets of the enzyme resulted in DMP323, which was studied in phase I clinical trials but found to suffer from variable pharmacokinetics in man. This report details the synthesis, conformational analysis, structure-activity relationships, and molecular recognition of this series of C2-symmetry HIV-1PR inhibitors. An initial series of cyclic ureas containing nonsymmetric P2/P2' is also discussed.
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PMID:Cyclic HIV protease inhibitors: synthesis, conformational analysis, P2/P2' structure-activity relationship, and molecular recognition of cyclic ureas. 878 49

Molecular models of HIV-1 protease and 21 peptide substrates with single amino acid substitutions at positions from P4 to P3' were built and compared with kinetic measurements. The crystal structure of HIV-1 protease with a peptidic inhibitor was modified to model the peptide substrate Pro-Ala-Val-Ser-Leu-Ala-Met-Thr for the starting geometry. Models were built of two reaction intermediates, HIV protease with peptide substrate and with its tetrahedral intermediate. The energy minimization used a new algorithm that increased the speed and eliminated a cut-off for non-bonded interactions. After minimization the models for substrate and tetrahedral intermediate both had root mean square deviations of 0.48 A for all atoms of the HIV protease compared to the starting crystal structure. Differences in the model structures and interaction energies for HIV protease with different substrates were analyzed. The calculated interaction energies for the 21 HIV protease-tetrahedral intermediate models gave a correlation coefficient of 0.64 with the kinetic measurements. The eight substrates with changes in the P1 and P1' residues next to the scissile bond gave the highest correlation of 0.93, while the 14 substrates with changes in P2-P2' gave a correlation coefficient of 0.86. The catalytic mechanism and factors influencing the catalytic efficiency of the different substrates are discussed in relation to the models. The predictive ability of molecular mechanics calculations is discussed in the context of the statistical mechanics analysis of the differences in free energy.
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PMID:Molecular mechanics calculations on HIV-1 protease with peptide substrates correlate with experimental data. 887 45

Several small, achiral nonpeptide inhibitors of HIV-1 protease with low micromolar activity were identified by mass screening of the Parke-Davis compound library. Two of the compounds, structurally similar, were both found to be competitive and reversible inhibitors [compound 1, 4-hydroxy-3-(3-phenoxypropyl)-1-benzopyran-2-one: Ki = 1.0 microM; compound 2, 4-hydroxy-6-phenyl-3-(phenylthio)-pyran-2-one: Ki = 1.1 microM]. These inhibitors were chosen as initial leads for optimization of in vitro inhibitory activity based on molecular modeling and X-ray crystallographic structural data. While improvements in inhibitory potency were small with analogues of compound 1, important X-ray crystallographic structural information of the enzyme-inhibitor complex was gained. When bound, 1 was found to displace H2O301 in the active site while hydrogen bonding to the catalytic Asps and Ile50 and Ile150. The pyranone group of compound 2 was found to bind at the active site in the same manner, with the 6-phenyl and the 3-phenylthio occupying P1 and P1', respectively. The structural information was used to develop design strategies to reach three or four of the internal pockets, P2-P2'. This work led to analogues of diverse structure with high potency (IC50 < 10 nM) that contain either one or no chiral centers and remain nonpeptide. The highly potent compounds possess less anti-HIV activity in cellular assays than expected, and current optimization now focuses on increasing cellular activity. The value of the HIV-1 protease inhibitors described is their potential as better pharmacological agents with a different pattern of viral resistance development, relative to the peptide inhibitors in human clinical trials.
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PMID:Discovery and optimization of nonpeptide HIV-1 protease inhibitors. 889 98

A combination of structure-activity studies, kinetic analysis, X-ray crystallographic analysis, and modeling were employed in the design of a novel series of HIV-1 protease (HIV PR) inhibitors. The crystal structure of a complex of HIV PR with SRSS-2,5-bis[N-(tert-butyloxycarbonyl)amino]-3,4-dihydroxy-1, 6-diphenylhexane (1) delineated a crucial water-mediated hydrogen bond between the tert-butyloxy group of the inhibitor and the amide hydrogen of Asp29 of the enzyme. Achiral, nonpeptidic 2-hydroxyphenylacetamide and 3-hydroxybenzamide groups were modeled as novel P2/P2' ligands to replace the crystallographic water molecules and to provide direct interactions with the NH groups of the Asp29/129 residues. Indeed, the symmetry-based inhibitors 7 and 19, possessing 3-hydroxy and 3-aminobenzamide, respectively, as a P2/P2' ligand, were potent inhibitors of HIV PR. The benzamides were superior in potency to the phenylacetamides and have four fewer rotatable bonds. An X-ray crystal structure of the HIV PR/7 complex at 2.1 A resolution revealed an asymmetric mode of binding, in which the 3-hydroxy group of the benzamide ring makes the predicted interaction with the backbone NH of Asp29 on one side of the active site only. An unexpected hydrogen bond with the Gly148 carbonyl group, resulting from rotation of the aromatic ring out of the amide plane, was observed on the other side. The inhibitory potencies of the benzamide compounds were found to be sensitive to the nature and position of substituents on the benzamide ring, and can be rationalized on the basis of the structure of the HIV PR/7 complex. These results partly confirm our initial hypothesis and suggest that optimal inhibitor designs should satisfy a requirement for providing polar interactions with Asp29 NH, and should minimize the conformational entropy loss on binding by reducing the number of freely rotatable bonds in inhibitors.
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PMID:Structure-based design of achiral, nonpeptidic hydroxybenzamide as a novel P2/P2' replacement for the symmetry-based HIV protease inhibitors. 889 4

The essential role of HIV-1 protease (HIV-1 PR) in the viral life cycle makes it an attractive target for the development of substrate-based inhibitors that may find efficacy as anti-AIDS drugs. However, resistance has arisen to potent peptidomimetic drugs necessitating the further development of novel chemical backbones for diversity based chemistry focused on probing the active site for inhibitor interactions and binding modes that evade protease resistance. AQ148 is a potent inhibitor of HIV-1 PR and represents a new class of transition state analogues incorporating an aminimide peptide isostere. A 3-D crystallographic structure of AQ148, a tetrapeptide isostere, has been determined in complex with its target HIV-1 PR to a resolution of 2.5 A and used to evaluate the specific structural determinants of AQ148 potency and to correlate structure-activity relationships within the class of related compounds. AQ148 is a competitive inhibitor of HIV-1 PR with a Ki value of 137 nM. Twenty-nine derivatives have been synthesized and chemical modifications have been made at the P1, P2, P1', and P2' sites. The atomic resolution structure of AQ148 bound to HIV-1 PR reveals both an inhibitor binding mode that closely resembles that of other peptidomimetic inhibitors and specific protein/inhibitor interactions that correlate with structure-activity relationships. The structure provides the basis for the design, synthesis and evaluation of the next generation of hydroxyethyl aminimide inhibitors. The aminimide peptide isostere is a scaffold with favorable biological properties well suited to both the combinatorial methods of peptidomimesis and the rational design of potent and specific substrate-based analogues.
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PMID:A new class of HIV-1 protease inhibitor: the crystallographic structure, inhibition and chemical synthesis of an aminimide peptide isostere. 889 11


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