Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UMLS:C0019693 (HIV)
 170,526 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Accelerated programmed cell death, or apoptosis, contributes to the CD4+ T-cell depletion characteristic of infection by human immunodeficiency virus (HIV). It has therefore been proposed that limiting apoptosis may represent a therapeutic modality for HIV infection. We found, however, that T leukemia cells or peripheral blood mononuclear cells (PBMCs) exposed to HIV-1 underwent enhanced viral replication in the presence of the cell death inhibitor, N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (z-AVD-fmk). Furthermore, z-VAD-fmk, which targets the pro-apoptotic interleukin-1 beta-converting enzyme (ICE)-like proteases, stimulated endogenous virus production in activated PBMCs derived from HIV-1-infected asymptomatic individuals. These findings suggest that programmed cell death may serve as a beneficial host mechanism to limit HIV spread and that strategies to inhibit it may have deleterious consequences for the infected host.
 ...
PMID:The inhibition of pro-apoptotic ICE-like proteases enhances HIV replication. 905 63

Two different crystal structures of the human immunodeficiency virus type 1 (HIV-1) integrase (IN) catalytic domain were analyzed for interactions at the enzyme active site. Gln-62 and Glu-92 interact with active-site residue Asp-64, and Lys-136 interacts with active-site residue Asp-116 across a dimer interface. Conservative and nonconservative substitutions were introduced at these positions to probe the roles of these interactions in HIV-1 integration. Purified mutant proteins were assayed for in vitro 3' processing, DNA strand transfer, and disintegration activities, and HIV-1 mutants were assayed for virion protein composition, reverse transcription, and infectivities in human cell lines. Each of the mutant IN proteins displayed wild-type disintegration activity, indicating that none of the interactions is essential for catalysis. Mutants carrying Gln or Ala for Glu-92 displayed wild-type activities, but substituting Lys for Glu-92 reduced in vitro 3' processing and DNA strand transfer activities 5- to 10-fold and yielded a replication-defective IN active-site mutant viral phenotype. Substituting Glu for Gln-62 reduced in vitro 3' processing and DNA strand transfer activities 5- to 10-fold without grossly affecting viral replication kinetics, suggesting that HIV-1 can replicate in T-cell lines with less than the wild-type level of IN activity. The relationship between IN solubility and HIV-1 replication was also investigated. We previously showed that substituting Lys for Phe-185 dramatically increased the solubility of recombinant IN but caused an HIV-1 particle assembly defect. Mutants carrying His at this position displayed increased solubility and wild-type replication kinetics, showing that increased IN solubility per se is not detrimental to virus growth.
 ...
PMID:Structure-based mutagenesis of the catalytic domain of human immunodeficiency virus type 1 integrase. 909 22

A new method for obtaining HIV-I protease was suggested. Fusion proteins composed of the N-terminal fragment of human gamma-interferon and HIV-I protease connected with (Asp)4Lys (protein I) or Asp-Pro (protein II) linkers were expressed in Escherichia coli cells. The fusion proteins were produced as insoluble inclusion bodies in the 20% yield of total cell protein. Protein I was cleaved by enterokinase. The solubility of protein I was increased by treating with Na-sulfite/Na-tetrathionate under denaturing conditions. Optimal conditions for efficient acidic hydrolysis of protein II at Asp-Pro bond were found. The hydrolysis products were separated by reversed-phase FPLC. The amount of tryptophan and cysteine residues in the enzyme obtained was estimated. The activity of HIV-I protease was determined using the chromogenic peptide. AlaArgVal NleNphGluAlaNleNH2 and a high-mol-wt substrate consisting of beta-galactosidase and a fragment of gag proteins, including p17-p24 processing site.
 ...
PMID:HIV-I protease. Cloning, expression, and purification. 910 Mar 48

To evaluate the available peptidic and pseudopeptidic inhibitors of HIV protease for their possible in vivo activity, a screening test using Escherichia coli was established. E. coli cells carrying the plasmid pET9c-PR containing the gene for HIV-1 protease under the control of a T7-promotor are grown in the absence and in the presence of inhibitors. The action of the toxic protease produced by the cells is counteracted by the inhibitors. Provided sufficient membrane permeability of the inhibitors exists, this results in accelerated cell growth. From the peptides known to be active in an in-vitro enzyme test, most compounds inhibit HIV protease only to a limited degree in this test. However, two short peptides (Ac-Ser-Tyr-Glu-Leu and Lys-Ile-Ser-Tyr-Asp-Tyr) protect cell growth to an considerabe extent, thus indicating that they reach the E. coli cytosol and there block HIV protease. Two pseudopeptides known to be very potent in the enzyme test (SDZ PRI 053 and CIBA 61755) also inhibit HIV-1 protease strongly in this cell growth test.
 ...
PMID:Screening of inhibitors of HIV-1 protease using an Escherichia coli cell assay. 914 91

Viral populations in a human immunodeficiency virus type 1 (HIV-1)-infected individual behave as a quasispecies with a rated distribution of fitness variants. Fitness distributions in naturally occurring viral populations have been difficult to study due to the lack of markers for individual virus clones and complicating inter- and intrahost factors like the presence of multiple cell types with distinct tropisms, differences in route of transmission, and intervening immunity. Here, we quantitated the relative fitness in vivo of three subpopulations of HIV-1 marked by mutations at codons 41 and 215 of reverse transcriptase (RT) directly related to zidovudine resistance in an untreated individual who was infected by a zidovudine-resistant strain transmitted from a donor on therapy. The transmission event did not have a substantial impact on the distribution of mutants within the dominant virus population replicating to high levels in the recipient. The evolution of the RT gene was monitored for 20 months. All 102 clones obtained from the donor and the recipient at the different time points contained the M41L mutation, which is associated with a fourfold reduction in zidovudine sensitivity. The leucine at position 41 was stable, although it was encoded by TTG and CTG triplets that fluctuated in abundance partially due to founder effects of clones with nonsilent mutations at codon 215. Of the three subpopulations in the patient, distinguished by a tyrosine (TAC), aspartic acid (GAC), or serine (TCC) at the 215 position of RT, the relative fitness of the GAC variant was calculated to be 10 to 25% higher than the initial TAC variant, and the relative fitness of the TCC variant was 1% higher than that of the GAC variant. Similar to other RNA viruses, lentivirus populations like HIV-1 in patients with a high virus load apparently consist of a broader spectrum of fitness variants than the 1 to 2% fitness difference sufficient for significant replicative advantage.
 ...
PMID:Broad spectrum of in vivo fitness of human immunodeficiency virus type 1 subpopulations differing at reverse transcriptase codons 41 and 215. 915 39

The chemokine receptor CCR5 is the major fusion coreceptor for macrophage-tropic strains of human immunodeficiency virus type 1 (HIV-1). To define the structures of CCR5 that can support envelope (Env)-mediated membrane fusion, we analyzed the activity of homologs, chimeras, and mutants of human CCR5 in a sensitive gene reporter cell-cell fusion assay. Simian, but not murine, homologs of CCR5 were fully active as HIV-1 fusion coreceptors. Chimeras between CCR5 and divergent chemokine receptors demonstrated the existence of two distinct regions of CCR5 that could be utilized for Env-mediated fusion, the amino-terminal domain and the extracellular loops. Dual-tropic Env proteins were particularly sensitive to alterations in the CCR5 amino-terminal domain, suggesting that this domain may play a pivotal role in the evolution of coreceptor usage in vivo. We identified individual residues in both functional regions, Asp-11, Lys-197, and Asp-276, that contribute to coreceptor function. Deletion of a highly conserved cytoplasmic motif rendered CCR5 incapable of signaling but did not abrogate its ability to function as a coreceptor, implying the independence of fusion and G-protein-mediated chemokine receptor signaling. Finally, we developed a novel monoclonal antibody to CCR5 to assist in future studies of CCR5 expression.
 ...
PMID:Two distinct CCR5 domains can mediate coreceptor usage by human immunodeficiency virus type 1. 926 47

The conformation of the DNA and the interactions of the nucleic acid with the protein in a complex of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) and 19-mer/18-mer double-stranded DNA template-primer (dsDNA) are described. The structure of this HIV-1 RT complex with dsDNA serves as a useful paradigm for studying aspects of nucleotide polymerases such as catalysis, fidelity, drug inhibition, and drug resistance. The bound dsDNA has a bend of approximately 41 degrees at the junction of an A-form region (first five base pairs near the polymerase active site) and a B-form region (the last nine base pairs toward the RNase H active site). The 41 degrees bend occurs smoothly over the four base pairs between the A-form portion and the B-form portion in the vicinity of helices alpha H and alpha I of the p66 thumb subdomain. The interactions between the dsDNA and protein primarily involve the sugar-phosphate backbone of the nucleic acid and structural elements of the palm, thumb, and RNase H of p66, and are not sequence specific. Amino acid residues from the polymerase active site region, including amino acid residues of the conserved Tyr-Met-Asp-Asp (YMDD) motif and the "primer grip," interact with 3'-terminal nucleotides of the primer strand and are involved in positioning the primer terminal nucleotide and its 3'-OH group at the polymerase active site. Amino acid residues of the "template grip" have close contacts with the template strand and aid in positioning the template strand near the polymerase active site. Helix alpha H of the p66 thumb is partly inserted into the minor groove of the dsDNA and helix alpha I is directly adjacent to the backbone of the template strand. Amino acid residues of beta 1', alpha A', alpha B', and the loop containing His539 of the RNase H domain interact with the primer strand of the dsDNA.
 ...
PMID:Protein-nucleic acid interactions and DNA conformation in a complex of human immunodeficiency virus type 1 reverse transcriptase with a double-stranded DNA template-primer. 935 57

Factors that modulate the placement of primer tRNA(3Lys) onto the viral RNA genome in human immunodeficiency virus type 1 (HIV-1) were investigated through analysis of reverse-transcribed products that are extended from the tRNA(3Lys) primer. Mutations were introduced into the HIV-1 pol gene to result in the appearance of a stop codon in the open reading frame of the reverse transcriptase (RT) gene. These constructs, BH10-RT1 and BH10-RT2, yielded viruses with truncated Pol proteins. Alternatively, we altered the sequences involved in frameshifting by generating the construct BH10-FS. With each of these mutated viruses, we found that the primer tRNA(3Lys) that was placed onto viral genomic RNA was present in an unextended state. In contrast, as expected, tRNA(3Lys) in the case of wild-type BH10 virus had been extended by 2 bases. Furthermore, the amount of tRNA(3Lys) that was placed onto viral RNA in mutated viruses was significantly less than that placed in the wild-type virus. We also generated a mutant within the polymerase-active site of RT (D185H) (Asp-->His) that eliminated RT polymerase activity. We found that the placement of primer tRNA(3Lys) onto viral genomic RNA was independent of enzyme function; however, the tRNA(3Lys) that was placed was present in an unextended state due to the loss of RT activity. In contrast, the elimination of protease activity through a D25A (Asp-->Ala) point mutation in the protease-active site (construct BH10-PR) did cause a drop in the efficiency of tRNA(3Lys) placement. In this situation, a proportion of the placed tRNA(3Lys) was found to be extended by 2 bases, although not to the extent found with wild-type virus (BH10), due to a decrease in RT activity associated with unprocessed Gag-Pol protein that could not be cleaved because of the loss of protease activity. We also investigated the role of the primer binding site (PBS) in the placement of tRNA(3Lys) through a series of 2-, 4-, and 8-nucleotide (nt) deletions at the 3' end of the PBS, i.e., BH10-PBS2, BH10-PBS4, and BH10-PBS8, respectively. In mutated viruses BH10-PBS2 and BH10-PBS4, the 2-base-extended form of tRNA(3Lys) was still detected. However, less primer tRNA(3Lys) was placed onto viral genomic RNA as more nucleotides were deleted until the percentage of placement seen with wild-type BH10 virus dropped to only 4% in the virus with 8 nt deleted (BH10-PBS8). Consistently, these mutated viruses possessed decreased initial replication capacity compared with that of the wild-type virus, with the extent of incapacity corresponding to the size of the deletion. However, after several days, an increase in replication potential was accompanied by a reversion to a wild-type PBS.
 ...
PMID:The roles of the human immunodeficiency virus type 1 Pol protein and the primer binding site in the placement of primer tRNA(3Lys) onto viral genomic RNA. 937 64

The CC-chemokine receptor CCR5 is required for the efficient fusion of macrophage (M)-tropic human immunodeficiency virus type 1 (HIV-1) strains with the plasma membrane of CD4+ cells and interacts directly with the viral surface glycoprotein gp120. Although receptor chimera studies have provided useful information, the domains of CCR5 that function for HIV-1 entry, including the site of gp120 interaction, have not been unambiguously identified. Here, we use site-directed, alanine-scanning mutagenesis of CCR5 to show that substitutions of the negatively charged aspartic acid residues at positions 2 and 11 (D2A and D11A) and a glutamic acid residue at position 18 (E18A), individually or in combination, impair or abolish CCR5-mediated HIV-1 entry for the ADA and JR-FL M-tropic strains and the DH123 dual-tropic strain. These mutations also impair Env-mediated membrane fusion and the gp120-CCR5 interaction. Of these three residues, only D11 is necessary for CC-chemokine-mediated inhibition of HIV-1 entry, which is, however, also dependent on other extracellular CCR5 residues. Thus, the gp120 and CC-chemokine binding sites on CCR5 are only partially overlapping, and the former site requires negatively charged residues in the amino-terminal CCR5 domain.
 ...
PMID:Amino-terminal substitutions in the CCR5 coreceptor impair gp120 binding and human immunodeficiency virus type 1 entry. 942 Feb 25

The cellular tropism of human immunodeficiency virus type 1 (HIV-1) is dependent on utilization of specific chemokine co-receptor: macrophage-tropic/non-syncytium-inducing (NSI) viruses use CCR5, whereas T-cell tropic/syncytium-inducing (SI) viruses preferentially use CXCR4. We have analyzed co-receptor usage of 24 phylogenetically distinct primary HIV-1 isolates representing group M (clades A-F) and group O with known SI and NSI phenotype, using lymphocytes from donor with nonfunctional CCR5 (CCR5-/-; homozygous 32-bp deletion). While all SI isolates infected CCR5-/- lymphocytes (and hence do not require CCR5 for viral entry), all NSI isolates, regardless of clade, did not infect CCR5-/- lymphocytes. Thus, CCR5 expression is required for infection with NSI isolates and the CCR5 usage is independent of viral genotype. To localize the viral determinant involved in CCR5 binding, the V3 sequences across the clades were aligned based on the CCR5 usage. There were conserved uncharged residues at position 11 of V3 (mostly serine/glycine) and negatively charged residues at residue 25 (mostly glutamic/aspartic acid) among all isolates that used CCR5, whereas substitution with arginine or glutamine at these two positions led to usage of a co-receptor other than CCR5. This analysis led us to identify a consensus motif S/GXXXGPGXXXXXXXE/D within the V3 loop that predicts CCR5 co-receptor usage. Most isolates, with exception of one isolate, containing the conserved motif and predicted to utilize CCR5 indeed had an absolute requirement of CCR5 expression for infectibility. Site-directed mutagenesis in the infectious molecular clone further confirmed these results. Taken together, these data provide evidence that sequences within the V3 loop provide important residues that might be directly or indirectly involved in binding to a CCR5 co-receptor.
 ...
PMID:CCR5 coreceptor usage of non-syncytium-inducing primary HIV-1 is independent of phylogenetically distinct global HIV-1 isolates: delineation of consensus motif in the V3 domain that predicts CCR-5 usage. 944 92


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>