Crystal structure of a complex of HIV‐1 protease with a dihydroxyethylene‐containing inhibitor: Comparisons with molecular modeling

Thanki, Narmada; Rao, Jayasimha; Foundling, S.; Howe, W. Jeffrey; Moon, Joseph B.; Hui, John O.; Tomasselli, A. G.; Heinrikson, Robert L.; Thaisrivongs, Suvit; Wlodawer, A.

doi:10.1002/pro.5560010811

Cited by 74 publications

(31 citation statements)

References 28 publications

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“…Although the peptides used for crystallization span a slightly different region of the cleavage site (P5 to P5Ј and P4 to P6Ј), these differences are likely to have minimal or no effect on the bound conformation of the substrate. From our previous experience with to determine substrate complexes with HIV protease (38,39,49), the P5 and P5Ј residues usually have no contact with the protease and are often disordered in the electron density. Thus, the presence or absence of P5 or P6Ј is unlikely to impact how the substrate is packed in the active site.…”

Section: Resultsmentioning

confidence: 99%

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Structural Basis for Coevolution of a Human Immunodeficiency Virus Type 1 Nucleocapsid-p1 Cleavage Site with a V82A Drug-Resistant Mutation in Viral Protease

Prabu-Jeyabalan

Nalivaika

King

et al. 2004

J Virol

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Maturation of human immunodeficiency virus (HIV) depends on the processing of Gag and Pol polyproteinsby the viral protease, making this enzyme a prime target for anti-HIV therapy. Among the protease substrates, the nucleocapsid-p1 (NC-p1) sequence is the least homologous, and its cleavage is the rate-determining step in viral maturation. In the other substrates of HIV-1 protease, P1 is usually either a hydrophobic or an aromatic residue, and P2 is usually a branched residue. NC-p1, however, contains Asn at P1 and Ala at P2. In response to the V82A drug-resistant protease mutation, the P2 alanine of NC-p1 mutates to valine (AP2V). To provide a structural rationale for HIV-1 protease binding to the NC-p1 cleavage site, we solved the crystal structures of inactive (D25N) WT and V82A HIV-1 proteases in complex with their respective WT and AP2V mutant NC-p1 substrates. Overall, the WT NC-p1 peptide binds HIV-1 protease less optimally than the AP2V mutant, as indicated by the presence of fewer hydrogen bonds and fewer van der Waals contacts. AlaP2 does not fill the P2 pocket completely; PheP1 makes van der Waals interactions with Val82 that are lost with the V82A protease mutation. This loss is compensated by the AP2V mutation, which reorients the peptide to a conformation more similar to that observed in other substrate-protease complexes. Thus, the mutant substrate not only binds the mutant protease more optimally but also reveals the interdependency between the P1 and P2 substrate sites. This structural interdependency results from coevolution of the substrate with the viral protease.Human immunodeficiency virus type 1 (HIV-1) matures after the viral protease processes (35) the Gag and Pol polyproteins at 10 substrate locations (3, 15). Therefore, inhibition of HIV-1 protease represents an important avenue for antiviral therapy (13, 48). The substrate sequences cleaved by the protease are nonhomologous (3), with the sequence of the nucleocapsid-p1 (NC-p1) substrate being the most different. In spite of the poor sequence homology among the substrate sites, a series of substrate-protease crystal structures led us to hypothesize that substrate specificity in HIV-1 protease results from the enzyme's recognizing an asymmetric shape (or envelope) rather than a particular amino acid sequence (40). This shape results from the conformation that a particular substrate sequence can adopt, implying that an interdependency necessarily exists among the different substrate residue sites.All of the protease inhibitors whose designs were structure based bind competitively (8,18,28,52,53) at the active site. Since these inhibitors bind at the same site as the substrates, many protease residues contact both substrates and inhibitors. Drug resistance, which often develops in the presence of therapeutic protease inhibitors, results from high viral turnover, the infidelity of the viral reverse transcriptase (16,42,43), and selective pressure on the virus. With drug resistance, the protease no longer binds as tightly to inhibitors but r...

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Section: Resultsmentioning

confidence: 99%

“…Crystals were grown by the hanging drop, vapor diffusion method as previously described (38,39,49). Stock solutions (25 mM) of substrate peptides used for cocrystallization were dissolved in refolding buffer rather than in dimethyl sulfoxide.…”

Section: Methodsmentioning

confidence: 99%

Structural Basis for Coevolution of a Human Immunodeficiency Virus Type 1 Nucleocapsid-p1 Cleavage Site with a V82A Drug-Resistant Mutation in Viral Protease

Prabu-Jeyabalan

Nalivaika

King

et al. 2004

J Virol

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“…Figure 3 generally represents the interacting mode of AHPBAs with HIVPR. Just like the inhibitor kni272 28 and most of the other transition state mimic inhibitors cocrystallized with HIVPR, 35,36 AHPBAs locate in the center of the typical binding pocket of HIVPR and share some common binding features for each other. All of the AHPBAs are bound in the active site of HIVPR in an extended conformation (Figures 1 and 2A), and the binding conformations of AHPBAs could be aligned quite well overall.…”

Section: Resultsmentioning

confidence: 99%

Elucidating the Inhibiting Mode of AHPBA Derivatives against HIV-1 Protease and Building Predictive 3D-QSAR Models

Huang

Luo

et al. 2001

J. Med. Chem.

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The Lamarckian genetic algorithm of AutoDock 3.0 has been used to dock 27 3(S)-amino-2(S)-hydroxyl-4-phenylbutanoic acids (AHPBAs) into the active site of HIV-1 protease (HIVPR). The binding mode was demonstrated in the aspects of the inhibitor's conformation, subsite interaction, and hydrogen bonding. The data of geometrical parameters (τ 1 , τ 2 , and τ 3 listed in Table 2) and root mean square deviation values as compared with the known inhibitor, kni272, 28 show that both kinds of inhibitors interact with HIVPR in a very similar way. The r 2 value of 0.860 indicates that the calculated binding free energies correlate well with the inhibitory activities. The structural and energetic differences in inhibitory potencies of AHPBAs were reasonably explored. Using the binding conformations of AHPBAs, consistent and highly predictive 3D-QSAR models were developed by performing CoMFA, CoMSIA, and HQSAR analyses. The reasonable r corss 2 values were 0.613, 0.530, and 0.717 for CoMFA, CoMSIA, and HQSAR models, respectively. The predictive ability of these models was validated by kni272 and a set of nine compounds that were not included in the training set. Mapping these models back to the topology of the active site of HIVPR leads to a better understanding of vital AHPBA-HIVPR interactions. Structural-based investigations and the final 3D-QSAR results provide clear guidelines and accurate activity predictions for novel HIVPR inhibitors.

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“…Crystals were obtained in hanging drops under more than one condition. The crystals used here were grown with a reservoir solution of 126 mM phosphate buffer (pH 6.2), 63 mM sodium citrate, and ammonium sulfate (28 to 31% and 22% for the substrates and inhibitors, respectively) (43,57). The crystals were brick shaped, tiny, and colorless, with a maximum dimension between 0.1 and 0.2 mm; the protein concentration ranged between 0.4 and 1.9 mg ml Ϫ1 .…”

Section: Methodsmentioning

confidence: 99%

Viability of a Drug-Resistant Human Immunodeficiency Virus Type 1 Protease Variant: Structural Insights for Better Antiviral Therapy

Prabu-Jeyabalan

Nalivaika

King

et al. 2003

J Virol

102

121

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Under the selective pressure of protease inhibitor therapy, patients infected with human immunodeficiency virus (HIV) often develop drug-resistant HIV strains. One of the first drug-resistant mutations to arise in the protease, particularly in patients receiving indinavir or ritonavir treatment, is V82A, which compromises the binding of these and other inhibitors but allows the virus to remain viable. To probe this drug resistance, we solved the crystal structures of three natural substrates and two commercial drugs in complex with an inactive drug-resistant mutant (D25N/V82A) HIV-1 protease. Through structural analysis and comparison of the protein-ligand interactions, we found that Val82 interacts more closely with the drugs than with the natural substrate peptides. The V82A mutation compromises these interactions with the drugs while not greatly affecting the substrate interactions, which is consistent with previously published kinetic data. Coupled with our earlier observations, these findings suggest that future inhibitor design may reduce the probability of the appearance of drug-resistant mutations by targeting residues that are essential for substrate recognition.

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Crystal structure of a complex of HIV‐1 protease with a dihydroxyethylene‐containing inhibitor: Comparisons with molecular modeling

Cited by 74 publications

References 28 publications

Structural Basis for Coevolution of a Human Immunodeficiency Virus Type 1 Nucleocapsid-p1 Cleavage Site with a V82A Drug-Resistant Mutation in Viral Protease

Structural Basis for Coevolution of a Human Immunodeficiency Virus Type 1 Nucleocapsid-p1 Cleavage Site with a V82A Drug-Resistant Mutation in Viral Protease

Elucidating the Inhibiting Mode of AHPBA Derivatives against HIV-1 Protease and Building Predictive 3D-QSAR Models

Viability of a Drug-Resistant Human Immunodeficiency Virus Type 1 Protease Variant: Structural Insights for Better Antiviral Therapy

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