Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: UNIPROT:Q9NRP7 (fused)
58,367 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To gain insights into the structure-function relationship of the envelope (env) glycoprotein of the human immunodeficiency virus type 1 (HIV-1) we have generated a vaccinia virus (VV) recombinant (VV-14kENV) that expresses a fusion protein (14k-env) consisting of the VV 14-kDa envelope protein (110 amino acids) fused at the C-terminus with HIV-1 env protein (816 amino acids). The 14k-env protein displayed unique structural properties in virus-infected cells. This protein was recognized by 14 kDa-specific antisera as well as HIV-1 env antisera. It was not cleaved during virus infection of cultured cells of various origins, it was stable, it was not released to the medium, and it was not incorporated into virions. Instead of a predicted 174-kDa protein, two proteins of about 110 and 100 kDa were observed. The size reduction of the fusion protein was due to limited glycosylation (110 kDa) and formation of unglycosylated protein (100 kDa). The 14k-env protein formed oligomeric structures and was exposed on the cell surface after virus infection. When mice were inoculated with the recombinant virus that expresses the 14K-env fusion protein, humoral immune response against gp160 was observed. Our findings suggest that 14k-env protein might display novel immunogenic properties.
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PMID:Structural properties of HIV-1 Env fused with the 14-kDa vaccinia virus envelope protein. 201 47

Stable transformants of the Jurkat T-cell line have been obtained that express either of two distinct forms of the type 1 human immunodeficiency virus nef gene: the nef-1-encoded protein (Nef-1) contains alanine, glycine, and valine at positions 15, 29, and 33, respectively; the protein specified by nef-2 (Nef-2) has threonine, arginine, and alanine at the corresponding positions. When Jurkat cells or their Nef-2-expressing transformants are treated with phorbol 12-myristate 13-acetate (PMA) plus either phytohemagglutinin (PHA) or antibodies against CD3 epsilon, T-cell receptor beta chain, or CD2, there is a prompt increase in interleukin 2 (IL-2) mRNA and intracellular calcium and in the IL-2 receptor alpha chain on the cell surface. Although cells expressing Nef-1 also induce calcium mobilization and the production of IL-2 receptor alpha chain, the formation of IL-2 mRNA is blocked in response to these stimuli. Moreover, Nef-1-expressing cells transfected with a plasmid in which the IL-2 promoter is fused to the chloramphenicol acetyltransferase (CAT) gene fail to induce CAT following treatment with PMA and PHA. By contrast, the parental and Nef-2-containing cells induce CAT normally. Nef-1-expressing cells can produce IL-2 mRNA in response to a combination of PMA and ionomycin, although much less efficiently than the parental Jurkat cells or Nef-2-expressing cells. These findings, and others described herein, suggest that the virally encoded Nef protein interferes with a signal emanating from the T-cell receptor complex that induces IL-2 gene transcription.
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PMID:Expression of the type 1 human immunodeficiency virus Nef protein in T cells prevents antigen receptor-mediated induction of interleukin 2 mRNA. 205 9

The human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein is essential for virus entry and the formation of multinucleated giant cells by cell fusion, one of the major virus-induced cytopathic effects. To study the effects of potential fusion inhibitors, a vaccinia virus recombinant expressing the envelope glycoprotein was generated and used to infect HeLa CD4+ cells. Syncytium induction was observed as early as 4 h postinfection and continued until the entire monolayer was fused. The N-terminus of the gp41 subunit of the HIV envelope protein is very hydrophobic, and appears to be involved in virus-induced membrane fusion. We synthesized several oligopeptide analogs of the N-terminal region of gp41 and determined their ability to inhibit HIV-induced cell fusion in CD4+ HeLa cells. A hexapeptide which was identical in amino acid sequence to the N-terminus of gp41 was found to completely inhibit cell fusion, whereas peptides with altered sequences showed reduced inhibitory activity. These peptides had no effect on protein synthesis, processing, or transport to the cell surface, and showed no signs of toxicity to cells even at very high concentrations. These results indicate that oligopeptides which are homologous to the fusion peptide of HIV inhibit virus-induced cytopathology, and should be evaluated further as potential antiviral agents.
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PMID:Oligopeptide inhibitors of HIV-induced syncytium formation. 207 10

Sequences encoding the human immunodeficiency virus type 1 (HIV-1) integrase gene have been cloned and expressed in Escherichia coli. The expressed protein is a lambda cII fusion protein of 37 kD containing the carboxyl-terminal 23 [corrected] amino acids of reverse transcriptase fused to the entire integrase sequence and is insoluble, a feature which allows partial purification away from soluble bacterial proteins. As judged by its reactivity with HIV positive sera in Western blot and in enzyme-linked immunosorbent assay (ELISA), the recombinant integrase retains antigenicity similar to native protein. Additionally, ELISA data obtained with the cloned protein indicate that patients infected with HIV-1 who are at different stages of progression to AIDS have antibodies reactive with the cloned integrase. HIV-2 positive human sera are also reactive with the cloned integrase. Rabbit antibodies produced against the recombinant protein react both by ELISA and Western blot with the homologous bacterially expressed protein, recognize both virion HIV-1 integrase and reverse transcriptase in Western blots, and immunoprecipitate an HIV-1 virion protein of 34 kD. Unlike human antisera from patients infected with HIV-1 or HIV-2 which are frequently reactive with both HIV-1 and HIV-2 integrase, the rabbit antibodies are type specific, reacting with HIV-1, but not with HIV-2 integrase by Western blot.
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PMID:Expression of HIV-1 integrase in E. coli: immunological analysis of the recombinant protein. 207 17

The rev gene of human immunodeficiency virus type 1 (HIV-1) encodes a 116 amino acid nuclear regulatory protein (Rev) that increases the cytoplasmic expression of viral mRNAs containing the Rev response element (RRE) and coding for the structural proteins, Gag and Env. To identify the functional domains of Rev, amino acid deletion and chain termination mutations were introduced in the Rev coding region. The ability of these mutants to increase the cytoplasmic expression of a Rev-test plasmid (pSV-AR), containing the RRE cloned into the 3' noncoding region of the CAT gene in plasmid pSV2CAT, was examined in transient expression assays in HeLa cells. Our results indicate that three distinct regions mapping within the N-terminal 98 amino acids of Rev are essential for its activity. The subcellular localization of the various Rev proteins was examined in COS cells by indirect immunofluorescence. Rev was found to localize predominantly in the nucleolus of transfected cells. All mutant Rev proteins, with the exception of a deletion mutant (rev delta 41-44) lacking four Arg residues of a highly basic domain, were found to localize in the nucleolus. Mutant rev delta 41-44 exhibited weak diffuse fluorescence in the nucleus with a tendency to accumulate in the cytoplasm. A 15 amino acid region encompassing this basic domain (38-52) when fused to the Escherichia coli beta-galactosidase gene efficiently directed the fusion gene product to the nucleus and nucleolus, suggesting a role for this domain in the nucleolar localization of Rev.
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PMID:Functional domains of the HIV-1 rev gene required for trans-regulation and subcellular localization. 210 12

The full-length envelope gene from an infectious human immunodeficiency virus type 2 (HIV-2) molecular clone was expressed in CD4+ and CD4- cells by a recombinant vaccinia virus vector. Pulse-chase experiments indicated that gp160 was processed into gp120 and gp41 subunits. Although large amounts of gp120 were shed into the medium, the recombinant vaccinia virus-infected cells fused with uninfected CD4+ cells. The receptor binding of HIV-2 gp120 was further analyzed using a panel composed of nine soluble CD4 mutants containing insertions of 2 amino acids within the first and second immunoglobulin-like domains. Of three mutations previously shown to interfere with HIV-1 gp120 binding, two also interfered with binding of the HIV-2 glycoprotein indicating use of the same binding site. Chemical crosslinking, sucrose gradient sedimentation, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis were employed to study the oligomerization of the envelope protein. The data indicated that gp160 assembles posttranslationally into dimers and higher oligomers that are probably tetramers.
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PMID:Synthesis, oligomerization, and biological activity of the human immunodeficiency virus type 2 envelope glycoprotein expressed by a recombinant vaccinia virus. 211 28

Translational effects of the RNA leader and Tat protein of human immunodeficiency virus type 1 (HIV-1) were investigated in rabbit reticulocyte lysate. Hybrid RNA species with natural or mutated HIV-1 leader fused to human interferon- gamma mRNA were produced in vitro from recombinant plasmids. HIV-1 leader RNA was found to inhibit translation through two mechanisms. A 3-fold trans-inhibition of translation was demonstrated by mixing hybrid HIV-1 leader RNA with indicator interferon mRNA. By comparison, HIV-1 leader caused a 50-fold cis-inhibition in lysate in which two trans-inhibitory factors, double-stranded RNA-dependent protein kinase and (2'-5')oligoadenylate synthetase, were suppressed. In contrast, purified HIV-1 Tat protein produced in Escherichia coli enhanced by 4-fold translation from HIV-1 leader-interferon mRNA but not from interferon mRNA lacking HIV sequences or from total poly(A)+ RNA. Translation of mRNA containing either a single base substitution in the loop of the "trans-acting responsive" sequence (TAR) or an alternative stem-loop in TAR was nevertheless stimulated by Tat. The enhancement of translation by Tat was largely due to relief of cis-inhibition, since the effect was found even in lysate in which double-stranded RNA-dependent protein kinase was inhibited with 2-aminopurine. These results suggest that translation is an important level of control in the replication cycle of HIV-1.
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PMID:Direct evidence for translational regulation by leader RNA and Tat protein of human immunodeficiency virus type 1. 212 Jul 1

FeLV-FAIDS, an immunodeficiency-inducing isolate of feline leukemia virus, is composed of a pathogenic but replication-defective genome (molecular clone 61C) and a replication-competent but non-immunodeficiency-inducing variant genome (molecular clone 61E). The chimeric virus EECC, composed of the 5' gag-pol of 61E fused to the env-3' LTR of 61C, also induces immunodeficiency. The 61C (or EECC) gp80 can be distinguished from that of 61E on the basis of antigenic recognition, size, and rate of posttranslational processing. We found that the nascent precursor polypeptides of the two viruses were the same size; however, the 61E gp80 rapidly shifted to a smaller size and was subsequently cleaved to gp70, whereas EECC gp80 maintained its nascent size and was cleaved to gp70 only after a prolonged time. Endo-beta-N-acetyl glucosaminidase H and N-glycanase digestions of newly formed glycoproteins resulted in a similar banding pattern for both viruses, indicating that both contained the same number of oligosaccharide side chains and that all of these were high mannose sugars. The metabolic inhibitors of glycosylation, castanospermine or N-methyldeoxynojirimycin, prevented both the rapid trimming of 61E gp80 and its cleavage to gp70. Treatment with mannosidase inhibitors, however, did not affect 61E gp80 processing or size, suggesting that retention of glucose residues on EECC was responsible for these distinguishing properties of the glycoprotein. The pathological consequence of aberrant viral glycoprotein processing was evaluated in feline 3201 T lymphocytes, which are infectable by both 61E and EECC but are killed only by EECC. As in fibroblasts, the EECC glycoprotein produced in lymphocytes was larger, antigenically distinct, and processed more slowly than was the glycoprotein of 61E. Castanospermine treatment of 61E-infected 3201 T cells, however, not only abrogated the antigenic differences between the 61E and EECC glycoproteins but also resulted in a cytopathic effect. Our results suggest that (i) intracellular accumulation of EECC envelope glycoprotein may occur consequent to retention of glucose residues on carbohydrate side chains and (ii) a strong correlation exists between delayed glycoprotein processing and cytopathicity in FeLV-FAIDS-infected T lymphocytes.
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PMID:Characterization and significance of delayed processing of the feline leukemia virus FeLV-FAIDS envelope glycoprotein. 216 20

Human hepatitis B virus (HBV) X-gene, previously shown to be capable of trans-activating heterologous regulatory elements of the human beta-interferon gene, the human immunodeficiency virus type I (HIV-1) long terminal repeat (LTR), the simian virus 40 (SV40), and HBV, has the capacity to code for a 17-kDa polypeptide (designated pX17). We now report that pX17 synthesized in Escherichia coli can activate transcription controlled by the HIV-1 LTR using a protoplast fusion technique. Protoplasts of E. coli-containing presynthesized X-protein were fused with lymphocytic H938 cells harboring an integrated copy of a plasmid with the CAT gene under control of the HIV-1 LTR (HIV-1 LTR CAT) and a marked increase in the steady state expression of the CAT mRNA was observed. When the same fused cells were treated with the protein synthesis inhibitor cyclohexamide, the pX17-dependent activation of the HIV-1 LTR was abolished. This result indicates that the X-protein expressed in E. coli is biologically active and suggests that the HBV X-protein-mediated trans-activation of the HIV-1 LTR in this system requires de novo cellular protein synthesis.
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PMID:Transcriptional activation of the human immunodeficiency virus type 1 long terminal repeat by hepatitis B virus X-protein requires de novo protein synthesis. 219

Different chimeric antibody-like molecules consisting of the four human CD4 extracellular domains (amino acids 1-369) fused to different parts of human IgG1 and IgM heavy-chain constant regions have been created and expressed in mammalian cells. For both IgG1 and IgM fusion proteins, the best expression in COS cells was observed for molecules lacking the CH1 domain of the heavy-chain constant region. The chimeric molecules are potent inhibitors of human immunodeficiency virus (HIV) infection and HIV-mediated cytotoxicity. A CD4:IgG1 hinge fusion protein, which was analyzed in more detail, binds efficiently to HIV gp160 and human Fc receptors and shows complement-assisted inhibition of viral propagation in culture. Half-life studies after intravenous application of the latter human fusion protein into mice and monkeys showed significant prolongation of serum survival compared to soluble CD4. An IgG2b murine homolog of the human CD4:IgG1 hinge fusion protein was prepared and evaluated in mice, where it was found to be nontoxic and to have no detectable effect on the humoral response to soluble antigen.
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PMID:Expression and characterization of human CD4:immunoglobulin fusion proteins. 219 3


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