Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Integrase”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 559 records · Page 31Linked to original sources
PubMed2003

Isolation, structure and HIV-1 integrase inhibitory activity of exophillic acid, a novel fungal metabolite from Exophiala pisciphila.

HIV-1 integrase is one of the three enzymes that are critical for replication and spread of HIV and its inhibition is one of the most promising new drug targets for anti-retroviral therapy with potential advantage over existing therapies. This paper describes the isolation and structure elucidation of exophillic acid, a novel dimeric 2,4-dihydroxy alkyl benzoic acid, derived from Exophiala pisciphila, a fungus isolated from a soil sample collected in Georgia, USA. Exophillic acid (1) and aquastatin A (2), a related compound, inhibited the strand transfer reaction of HIV-1 integrase with IC50 values of 68 and 50 microM, respectively.

Benzoates↗
PubMed1999

Analysis of integration activity of human immunodeficiency virus type-1 integrase.

The integration activity of human immunodeficiency virus type-1 (HIV-1) integrase was characterized in vitro by using pre-processed oligonucleotide substrates. The highest level of integration activity was found at pH 6.5 to 7.0, while the endonucleolytic activity was highest at pH 7.4 to 8.0. Although the endonucleolytic and integration reactions are consecutive in retroviral integration, our result indicates that the optimal conditions of the two reactions are quite different. In addition, it is suggested that the endonucleolytic and integration steps can be separated by control of the cellular physiological state in retroviral therapy. Strong integration was detected in the presence of 0.5-10 mM Mn2+ ion, but weak integration at around 10 mM Mg2+ ion. This observation explains that the Mn2+ ion is preferred to the Mg2+ ion as a cofactor in the integration reaction. Although there was no sequence-specificity in the integration site of the target DNA, integration was found to frequently occur at particular regions of the target DNA. Furthermore, the mutant integrases such as Asp116, Ser147, and Glu152, which had been reported previously, were shown to lose integration activity completely, indicating that these residues are critically involved in catalytic action.

Base Sequence↗
PubMedDate not supplied

Inhibition of HIV-1 replication and infectivity by expression of a fusion protein, VPR-anti-integrase single-chain variable fragment (SFv): intravirion molecular therapies.

OBJECTIVES: To deliver antiretroviral agents or other foreign proteins into progeny virions and evaluate their inhibitory effect on human immunodeficiency virus type 1 (HIV-1) replication. STUDY DESIGN/METHODS: HIV-1 encodes proteins in addition to gag, pol, and env, some of which are packaged into virus particles. One essential retroviral enzyme is integrase (IN), which has been used as a target for developing agents that inhibit virus replication. In previous studies, we demonstrated that intracellular expression of single-chain variable antibody fragments (SFvs), which bind to IN, results in resistance to productive HIV-1 infection in T-lymphocytic cells. Because the highly conserved accessory HIV-1 Vpr protein can be packaged within virions in quantities similar to those of the major structural proteins, this primate lentiviral protein may be used as a fusion partner to deliver antiviral agents or other foreign proteins into progeny virions. In these studies, the fusion proteins Vpr-chloramphenicol acetyl transferase (CAT) and Vpr-SFv-IN have been developed. Stable transfectants expressing these fusion proteins were generated from PA317 cells and SupT1 T-lymphocytic cells and analyzed using immunofluorescence microscopy. After challenge of SupT1 cells with HIV-1, p24 antigen expression was evaluated. The incorporation of these fusion proteins were evaluated by immunoprecipitation of virions using a Vpr antibody. RESULTS: Expression of the fusion proteins was confirmed by immunofluorescent staining in PA317 cells transfected with the plasmids expressing Vpr-CAT and Vpr-SFv-IN proteins. Stable transfectants expressing these fusion proteins were generated from SupT1 T-lymphocytic cells. When challenged, HIV-1 replication, as measured by HIV-1 p24 antigen expression, was inhibited in cells expressing Vpr-SFv-IN. It was demonstrated that Vpr-chloramphenicol acetyl transferase (Vpr-CAT and Vpr-SFv-IN proteins can be efficiently packaged into the virions and that Vpr-SFv-IN also decreases the infectivity of virions into which it is encapsidated. CONCLUSIONS: An anti-integrase single-chain variable fragment moiety can be delivered into HIV-1 virions by fusing it to Vpr. Vpr-SFv-IN decreases HIV-1 production in human T-lymphocytic cells. The benefits of "intravirion" gene therapy include immunization of target cells as well as decreasing infectivity of HIV-1 virions harboring the fusion construct. Thus, this approach to anti-HIV-1 molecular therapies has the potential to increase inhibitory effects against HIV-1 replication and virion spread.

Anti-HIV Agents↗
PubMed2004

[Cloning the gene fragment coding the putative integrase-like protein from X maltophilia].

OBJECTIVE: To amplify the nucleotide sequence of CG-like receptor from X anthomonas maltophilia(X maltophilia). METHODS: Using the specific primer P1 designed by Grover's reported 342 bp partial nucleotide sequence of X maltophilia CG-like receptor and random primer to PCR amplify, PCR product was cloned in the pUCm-T vector. After the recombinant plasmid was tested by restriction endonuclease digestion, the insert on the recombinant plasmid was sequenced and analyzed. RESULTS: About 500 bp PCR product was cloned in the pUCm-T vector and obtained the recombinant pUCm-Int. By sequencing to the insert on the pUCm-Int with M13 universal sequencing primers, the 410-486 bp fragment of the cloned 510 bp nucleotide sequence (GenBank accession number: AY363962) showed 84% identity with the 9304-8958 bp fragment of the XACb0009 gene on plasmid pXAC64 of Xanthomonas axonopodis pv. citri. And the 4-166aa fragment of its translated 169aa sequence had 62% identity with the 38-200aa sequence of integrase-like protein coded by the XACb0009 gene. CONCLUSION: The cloned 510 bp nucleotide sequence was possibly the partial gene sequence coding the integrase-like protein of X maltophilia.

Amino Acid Sequence↗
PubMed2006

MK-0518 and GS-9137: two promising integrase inhibitors in the pipeline.

Possibly the most exciting news to come out of the 13th Conference on Retroviruses and Opportunistic Infections, held in Denver last February, concerned clinical trials of two experimental drugs in a new class: integrase inhibitors. If successful in further trials, integrase inhibitors could revitalize the treatment regimens of people living with multidrug-resistant HIV.

Drug Resistance, Viral↗
PubMed1997

Characterization of human immunodeficiency virus type 1 integrase mutants expressed in Escherichia coli.

The eight mutant integrase (IN) proteins of human immunodeficiency virus type (HIV-1), which have a single point mutation at a highly conserved central region, were prepared, and characterized in terms of their endonucleolytic activities and disintegration activities in vitro. Mutation of two highly conserved amino acids, Asp116 or Glu152, leads to complete loss of both the activities, suggesting that these two amino acids are directly associated with enzymatic functions. In addition, the mutant of the position Ser147 was found to have highly depressed endonucleolytic activity showing that the reaction was very delayed in comparison with that of the wild type. However, significant disintegration was detected in the mutant Ser147, indicating that the enzymatic mechanisms of the endonucleolytic and disintegration activities are not exactly reverse. The integrase protein with a mutation at the conserved amino acid Asn117 or Gly118 had a slight loss of the endonucleolytic activity, while a mutation at the three positions, Tyr143, Ser153, and Lys159, had no detectable effect on their enzymatic activities. These results indicate that only a few of the conserved amino acids are critical for enzymatic activities.

Amino Acid Sequence↗
PubMed2005

Identification of amino acids in HIV-1 and avian sarcoma virus integrase subsites required for specific recognition of the long terminal repeat Ends.

A tetramer model for HIV-1 integrase (IN) with DNA representing 20 bp of the U3 and U5 long terminal repeats (LTR) termini was assembled using structural and biochemical data and molecular dynamics simulations. It predicted amino acid residues on the enzyme surface that can interact with the LTR termini. A separate structural alignment of HIV-1, simian sarcoma virus (SIV), and avian sarcoma virus (ASV) INs predicted which of these residues were unique. To determine whether these residues were responsible for specific recognition of the LTR termini, the amino acids from ASV IN were substituted into the structurally equivalent positions of HIV-1 IN, and the ability of the chimeras to 3 ' process U5 HIV-1 or ASV duplex oligos was determined. This analysis demonstrated that there are multiple amino acid contacts with the LTRs and that substitution of ASV IN amino acids at many of the analogous positions in HIV-1 IN conferred partial ability to cleave ASV substrates with a concomitant loss in the ability to cleave the homologous HIV-1 substrate. HIV-1 IN residues that changed specificity include Val(72), Ser(153), Lys(160)-Ile(161), Gly(163)-Val(165), and His(171)-Leu(172). Because a chimera that combines several of these substitutions showed a specificity of cleavage of the U5 ASV substrate closer to wild type ASV IN compared with chimeras with individual amino acid substitutions, it appears that the sum of the IN interactions with the LTRs determines the specificity. Finally, residues Ser(153) and Val(72) in HIV-1 IN are among those that change in enzymes that develop resistance to naphthyridine carboxamide- and diketo acid-related inhibitors in cells. Thus, amino acid residues involved in recognition of the LTRs are among these positions that change in development of drug resistance.

Amino Acid Sequence↗
PubMed2005

Comparison of multiple molecular dynamics trajectories calculated for the drug-resistant HIV-1 integrase T66I/M154I catalytic domain.

HIV-1 integrase (IN) is an essential enzyme for the viral replication and an interesting target for the design of new pharmaceuticals for multidrug therapy of AIDS. Single and multiple mutations of IN at residues T66, S153, or M154 confer degrees of resistance to several inhibitors that prevent the enzyme from performing its normal strand transfer activity. Four different conformations of IN were chosen from a prior molecular dynamics (MD) simulation on the modeled IN T66I/M154I catalytic core domain as starting points for additional MD studies. The aim of this article is to understand the dynamic features that may play roles in the catalytic activity of the double mutant enzyme in the absence of any inhibitor. Moreover, we want to verify the influence of using different starting points on the MD trajectories and associated dynamical properties. By comparison of the trajectories obtained from these MD simulations we have demonstrated that the starting point does not affect the conformational space explored by this protein and that the time of the simulation is long enough to achieve convergence for this system.

Biophysics↗
PubMed2006

Discovery and structure-activity relationship studies of a unique class of HIV-1 integrase inhibitors.

HIV-1 integrase (IN) is an essential enzyme for viral replication and a validated target for the development of drugs against AIDS. Currently there are no approved drugs that target IN. However, new IN inhibitors are under clinical investigation. As more IN inhibitors enter human drug trials, there is a growing need for the design of novel lead compounds with diverse structural scaffolds and promising pharmacokinetic properties to counteract the difficulties observed with first-generation IN inhibitors. We have identified a novel class of IN inhibitors through the systematic exploration of structure-activity relationships in a series of linomide analogues. The predicted bound conformation of the most active analogues inside the IN active site also supports the observed structure-activity correlation in this new compound class.

Base Sequence↗
PubMed2006

Inhibitory activity on HIV-1 reverse transcriptase and integrase of a carmalol derivative from a brown Alga, Ishige okamurae.

The bioassay-directed isolation of a marine brown alga, Ishige okamurae, afforded a carmalol derivative, diphlorethohydroxycarmalol. This compound exhibited inhibitory effects on HIV-1 reverse transcriptase and integrase with IC(50) values of 9.1 microM and 25.2 microM, respectively. However, diphlorethohydroxycarmalol did not show an inhibitory activity against HIV-1 protease. Moreover, diphlorethohydroxycarmalol nonaacetate obtained by acetylation and fucosterol failed to show any inhibitory activity against these viral enzymes.

Anti-HIV Agents↗
PubMed2000

Cellular distribution and karyophilic properties of matrix, integrase, and Vpr proteins from the human and simian immunodeficiency viruses.

Infections by human and simian immunodeficiency viruses (HIV and SIV) are independent of host cell division since the preintegration complex (PIC), containing the viral DNA, is able to undergo active nuclear import after viral entry. In order to clarify the mechanisms responsible for nuclear import of the PIC, we have analyzed the subcellular distribution and the karyophilic properties of its viral components, matrix protein (MA), integrase (IN), Vpr, and Vpx. Although MA has been reported to contain a nuclear localization signal, the MA/GFP fusions are excluded from the nucleus and associated with cellular membranes. In contrast, both HIV-1 and SIV IN and Vpr localize in the nucleus of transfected cells. Interestingly, only Vpx from SIVsm virus accumulate in the nucleus while SIVsm Vpr is uniformly distributed throughout nucleus and cytoplasm. Coexpression of MA, Vpr, and IN does not induce any change in their respective intracellular localizations. Finally, we confirm the karyophilic properties of HIV-1 IN and Vpr using an in vitro nuclear import assay. These results indicate that the viral proteins IN and Vpr, which are strongly associated with the viral DNA within PIC, may participate in the nuclear import of the HIV PIC.

Animals↗
PubMed2002

Analysis of conserved and non-conserved amino acids critical for ALSV (Avian leukemia and sarcoma viruses) integrase functions in vitro.

Retroviral integrase (IN) is the viral enzyme responsible for the integration of viral DNA into host cellular DNA. In vitro, recombinant IN protein is able to catalyze the 3'-processing, strand transfer and disintegration activities. In order to analyze the importance of specific residues of ALSV (Avian leukemia and sarcoma viruses) IN protein, we introduced 31 amino acid substitutions either in residues previously shown by others to be involved in IN oligomerization or in selected conserved and non-conserved residues through the IN sequence. We tested, in vitro, the three catalytic activities of these mutants as well as their capacity to bind DNA. We found that (i) 88% of the substitutions occurring on well-conserved residues have an effect on IN activities (ii) two mutants (S85T in the central catalytic domain and N197C in the C-terminal domain) present a reduced efficiency of DNA binding compared to the wild type protein. Moreover, all mutations made on the dimer interface of C-terminal domain present reduced activities, suggesting an important role of this part of the protein. Finally, for some mutations, we observed differences between the ALSV and HIV (Human immunodeficiency virus) IN corresponding residues.

Alpharetrovirus↗
PubMed2004

The use of a new in vitro reaction substrate reproducing both U3 and U5 regions of the HIV-1 3'-ends increases the correlation between the in vitro and in vivo effects of the HIV-1 integrase inhibitors.

Human Immunodeficiency Virus type 1 (HIV-1) integrase (IN) is an attractive target for the development of new antiviral therapies. Recently, several HIV-1 recombinant IN (rIN) in vitro inhibitors have been described. However, the great majority of them failed to block the virus replication in cell-based assays, suggesting the inadequacy of the in vitro assay systems used for inhibitor screening. To improve these systems, we designed a 40(mer) duplex DNA reaction substrate consisting of recognition sequences from both U3 and U5 HIV-1 long terminal repeat (LTR) termini. The HIV-1 rIN was able to catalyze its enzyme activities recognizing both ends of the 40(mer) dsDNA. Using this substrate we assayed the effects on rIN catalysis of different classes of compounds which inhibit the HIV-1 rIN in vitro when the reaction substrate is the standard 21(mer) U5 dsDNA, and that are either active or inactive on the HIV-1 replication. We also compared the efficacy of these compounds when added to the reaction before or after the formation of the rIN-dsDNA complex. In this system, the enzyme preincubation with the two-ended 40(mer) dsDNA before the addition of the compounds allowed a strong correlation between the effects of hydroxylated aromatics derivatives on rIN activity in cell-free assays and their effects on viral replication in cell-culture assays. This increase in drug selectivity of the rIN in vitro assay was explored by investigating whether it was due to the length of the 40(mer), longer than the standard 21(mer), or to presence of both viral ends, versus only one viral end. To this purpose we designed four 40(mer) oligonucleotides containing either only one viral end or two-repetitive ends, finding that the architecture of the rIN-dsDNA complex and its compound susceptibility is significantly influenced by the sequence of the dsDNA substrate.

Anti-HIV Agents↗
PubMed2004

Synthesis, antiviral, and anti-HIV-1 integrase activities of 3-aroyl-1,1-dioxo-1,4,2-benzodithiazines.

HIV-1 integrase (IN) is an essential enzyme for effective viral replication and is an attractive target for selective blockade of viral infection. Previously, we identified a series of sulfones, sulfonamides, and mercaptosalicylhydrazides (MBSAs) as IN inhibitors with antiviral activities in cell-based assays. In an effort to optimize a series of our active site directed lead compounds, we designed and synthesized novel benzodithiazines starting from MBSAs. In contrast to all reported IN inhibitors benzodithiazines are essentially nontoxic. Significant antiviral potency was only observed at concentration exceedingly higher than that required to inhibit purified IN.

Anti-HIV Agents↗
PubMed2004

Active site binding modes of the beta-diketoacids: a multi-active site approach in HIV-1 integrase inhibitor design.

Predicting a bioactive conformation of a ligand is of paramount importance in rational drug design. The task becomes very difficult when the receptor site possesses a region with unusual conformational flexibility. Significant conformational differences are present in the active site regions in the available crystal structures of the core domains of HIV-1 integrase (IN). Among all reported IN inhibitors, the beta-diketoacid class of compounds has proved to be of most promise and indeed S-1360 was the first IN inhibitor to enter clinical studies. With an aim to predict the bioactive (active site bound) conformation of S-1360, we performed extensive docking studies using three different reported crystal structures where the active site or partial active site region was resolved. For comparison we extended our studies to include 5CITEP (the first compound cocrystallized with IN core domain) and a bis-diketoacid (BDKA). We found that the conformation of S-1360 when bound in one of the active sites matches that of the experimentally observed results of IN escape mutants resistant to S-1360. Therefore, we propose that this active site conformation is the biologically relevant conformation and can be used for the future structure-based drug design studies selectively targeting IN.

Anti-HIV Agents↗
PubMed2006

Mining the NCI antiviral compounds for HIV-1 integrase inhibitors.

HIV-1 integrase (IN) is an essential enzyme for effective viral replication and is a validated target for the development of antiretroviral drugs. Currently, there are no approved drugs targeting this enzyme. In this study, we have identified 11 structurally diverse small-molecule inhibitors of IN. These compounds have been selected by mining the moderately active antiviral molecules from a collection of 90,000 compounds screened by the National Cancer Institute (NCI) Antiviral Program. These compounds, which were screened at the NCI during the past 20 years, resulted in approximately 4000 compounds labeled as 'moderately active.' In our study, chalcone 11 shows the most potent activity with an IC(50) of 2+/-1 microM against purified IN in the presence of both Mn(2+) and Mg(2+) as cofactors. Docking simulations using the 11 identified inhibitors as a training set have elucidated two unique binding areas within the active site: the first encompasses the conserved D64-D116-E152 motif, while the other involves the flexible loop region formed by amino acid residues 140-149. The tested inhibitors exhibit favorable interactions with important amino acid residues through van der Waals and H-bonding contacts.

Anti-HIV Agents↗
PubMed2006

Biotinylated biphenyl ketone-containing 2,4-dioxobutanoic acids designed as HIV-1 integrase photoaffinity ligands.

The diketo acid (DKA) class of HIV-1 integrase inhibitors are thought to function by chelating divalent metal ions within the enzyme catalytic center. However, differences in mutations conferring resistance among sub-families of DKA inhibitors suggest that multiple binding orientations may exist. In order to facilitate identification of DKA-binding sites, biotin-tagged biphenyl ketone-containing 2,4-dioxobutanoic acids were prepared as DKA photoaffinity probes. Introduction of biotin was obtained by means of Huisgen [3+2] cycloaddition 'click chemistry.' Two photoprobes, 5a and 5b, were prepared bearing short and long linker segments, respectively, between the biotin and DKA nucleus. The greatest inhibitory potency was shown by 5b, which inhibited 3'-processing and strand transfer reactions with IC50 values of > 333 microM and 12.4 microM, respectively. In cross-linking assays designed to measure disruption of substrate DNA binding, the photoprobes behaved similarly to a reference DKA inhibitor. Analogues 5a and 5b represent novel photoaffinity ligands, which may be useful in clarifying the HIV-1 binding interactions of DKA inhibitors.

Acetoacetates↗
PubMed2004

HIV-1 integrase pharmacophore model derived from diverse classes of inhibitors.

A three-dimensional pharmacophore model has been generated for HIV-1 integrase (HIV-1 IN) from known inhibitors. A dataset consisting of 26 inhibitors was selected on the basis of the information content of the structures and activity data as required by the catalyst/HypoGen program. Our model was able to predict the activity of other known HIV-1 IN inhibitors not included in the model generation, and can be further used to identify structurally diverse compounds with desired biological activity by virtual screening.

HIV Integrase↗