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: UMLS:C0021051 (immunodeficiency)
71,517 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Recombinant vaccinia viruses containing either the entire gag/pol gene or the reverse transcriptase (RT) domain of the human immunodeficiency virus (HIV) were constructed. In mammalian cells infected with the recombinant vaccinia virus containing the gag/pol gene, major and minor polypeptides of 55 and 41 kDa were made, but processed gag products (p24/p17/p15) were not detected. In addition, none of the products of the pol open-reading frame were seen. Both the 55- and 41-kDa gag proteins were post-translationally modified by addition of myristic acid residues in recombinant vaccinia-infected cells, and were immunoprecipitated by antiserum to p24 gag, as well as by antisera from HIV-infected patients. These results indicate that neither proteolytic processing nor other HIV proteins are required for myristilation, and suggest that the 55- and 41-kDa gag precursors share the same amino terminus as p17. Cells infected with a separate vaccinia recombinant containing a truncated piece of the gag/pol gene with added start and stop codons at the 5' and 3' ends of the RT reading frame synthesized a major 61-kDa and a minor 51-kDa protein product which reacted immunologically with both a monoclonal antibody to native HIV p66/51 and antisera from HIV-infected patients. These proteins were purified from recombinant vaccinia-infected mammalian cells, and their enzyme activity was found to be similar to that of authentic HIV RT. Cells infected with the vaccinia/RT vector contained approximately 200-fold more RT per milligram of protein than cells infected with HIV. Recombinant RT was inhibited by dideoxynucleoside triphosphates and should be useful in screening for specific inhibitors of this enzyme. Mice inoculated intradermally with 10(8) plaque-forming units of the vaccinia/RT vector developed specific antibodies to the p66/51 proteins of HIV, but anti-HIV antibodies were not detected in mice inoculated with the vaccinia/gag vector.
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PMID:Characterization of human immunodeficiency virus gag/pol gene products expressed by recombinant vaccinia viruses. 245 42

The proteins p15 and p24 of the human immunodeficiency virus (HIV) type 1 gag gene were expressed as fusion proteins in Escherichia coli for detecting antibodies against the acquired immunodeficiency virus by Western blot (immunoblot) analysis. These fusion proteins contain amino acids 1 to 375 of the E. coli beta-galactosidase linked to the viral protein(s) by a recognition sequence for the specific protease factor Xa. They are obtained in large amounts in insoluble inclusion bodies. To avoid ambiguous results caused by cross-reaction of sera with bacterial proteins in Western blots, we purified the recombinant fusion proteins and subsequently removed the bacterial part of the fusions by cleavage with factor Xa. The cleavage mixtures were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and blotted onto nitrocellulose membranes. The viral proteins obtained by this method did not contain any bacterial proteins or protein fragments. Thus, false-positive results in HIV Western blot analysis with bacterially expressed HIV proteins can be excluded with these purified recombinant viral antigens.
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PMID:Cleavage of procaryotically expressed human immunodeficiency virus fusion proteins by factor Xa and application in western blot (immunoblot) assays. 250 57

The virally encoded protease of human immunodeficiency virus is responsible for the processing of the gag and gag-pol polyprotein precursors to their mature polypeptides. Since correct processing of the viral polypeptides is essential for the production of infectious virus, HIV protease represents a potential target for therapeutic agents that may prove beneficial in the treatment of AIDS. In this study, full-length gag polyprotein has been synthesized in vitro to serve as a substrate for bacterially expressed HIV-1 protease. Expression of the protease in E. coli from the lac promoter was enhanced approximately five-fold by deletion of a potential hairpin loop upstream from the codon determining the amino terminus of mature protease. Extracts of induced cultures of E. coli harboring a protease-containing plasmid served as the source of protease activity. The gag polyprotein synthesized in vitro was cleaved by such lysates, producing fragments corresponding in size to p17 plus p24 and mature p24. Immunoprecipitations with monoclonal antibodies to p17 and p24 polypeptides suggest that initial cleavage of gag polyprotein occurs near the p24-p15 junction. The proteolysis was inhibited by pepstatin with an IC50 of 0.15 mM for cleavage at the p24-p15 junction and 0.02 mM for cleavage at the p17-p24 junction.
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PMID:Cleavage of HIV-1 gag polyprotein synthesized in vitro: sequential cleavage by the viral protease. 269 58

Human T-cell leukemia virus (HTLV) type I-related endogenous sequences (HRES) have been cloned from a human genomic library. HRES-1/1 is present in DNA of all normal donors examined. By nucleotide sequence analysis, HRES-1/1 contains two potential open reading frames capable of encoding a p25 and a p15. A 684 bp flanking region 5' from the first ATG codon of p25 contains a TATA-box, a poly-adenylation signal, a putative tRNA primer binding site, and inverted repeats at locations which are typical of a retroviral long terminal repeat. Phylogenetic analysis suggests that HRES-1/1 entered the genome in primates, presumably as an exogenous retrovirus. From the deduced amino acid sequence of HRES-1/1 p25, residues 6-36 show a sequence homology of 32% and 39% to gag region segments of HTLV-I and HTLV-II, while residues 104-139 display a sequence homology of 33% and 28% to the gag regions of human immunodeficiency virus type 2 (HIV-2) and feline sarcoma virus (FSV), respectively. This suggests that the original exogenous virus infecting primate may be chimeric in structure. The HRES-1/1 genomic locus is transcriptionally active in lymphoid cells, melanoma cells, and embryonic tissues.
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PMID:Detection and cloning of new HTLV-related endogenous sequences in man. 278 Mar 12

We have used a recombinant vaccinia virus (VV) which expresses high levels of human immunodeficiency virus-1 (HIV-1) gag proteins to analyze the processing pathway of the gag p55 precursor. HIV-1 gag proteins were isolated from [3H]leucine-labeled VV:gag-infected H9 T lymphocytes by immunoprecipitation with either anti-p24, anti-p17, or anti-p6 antibodies. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed that processing of the p55 precursor involves three major intermediates (p41a, p41b, and p39). The p41a and p39 proteins contain the p17 and p24 protein segments, and the p41b is comprised of p24 and p15 segments. On two-dimensional gels, each intermediate as well as the mature p24 and p17 proteins migrated as distinct species. [3H]Myristic acid labeling of the HIV-1 gag proteins revealed that in addition to p55 and p17, the p41a and p39 intermediates, but not p41b, are myristylated, confirming that myristylation occurs at the NH2 terminus before cleavage of the p55 precursor protein. We conclude that the myristylated HIV-1 gag p55 precursor is initially cleaved at random either at the p17/p24 junction or at two sites between p24 and p15 proteins, resulting in three intermediates (p41a, p41b, and p39) which are subsequently cleaved to yield mature gag proteins.
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PMID:Identification of protein intermediates in the processing of the p55 HIV-1 gag precursor in cells infected with recombinant vaccinia virus. 278 91

We describe here a one step HPLC technique for purifying the four gag proteins (p26, p15, p11 and p9) and two env glycoproteins (gp90 and gp45) from purified equine infectious anemia virus (EIAV), a member of the lentivirus subfamily of retroviruses. The purification procedure employs a reverse-phase phenyl Radial-pak cartridge contained in a high pressure radial compression chamber in which a shallow, multistep acetonitrile gradient is applied at ambient temperatures. The purified proteins are recovered at an efficiency of 60-70%. Moreover, the isolated components retain their antigenicity and are suitable for a variety of biochemical analyses including protein sequencing. The purification of EIAV gp90 and gp45 represents the first successful isolation of a lentivirus glycoprotein from purified virus preparations. The availability of these separated proteins permitted direct protein sequencing which confirmed the previously reported env gene sequence and provides important antigens for the development of diagnostic immunoassays and subunit vaccines. The procedures described appear applicable to other lentiviruses, including human immunodeficiency virus (HIV), and perhaps to hydrophobic membrane proteins in general.
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PMID:Lentivirus antigen purification and characterization: isolation of equine infectious anemia virus gag and env proteins in one step by reverse phase HPLC and application to human immunodeficiency virus glycoproteins. 283 62

Antibodies to specific HIV viral antigens were measured by ELISA recombinant proteins representing gag and env amino acid sequences of human immunodeficiency virus (HIV) (E. I. du Pont de Nemours, Wilmington, DE) and by a Western blot system using biotinavidin detection (Biotech Research Labs, Rockville, MD) on 36 HIV antibody-positive hemophiliacs (HTLV-III ELISA, du Pont) on whom date of seroconversion was known and on whom serial samples where available between 1977 and 1986, representing 2-8 years following seroconversion. The 36 included 9 acquired immunodeficiency syndrome (AIDS) and 27 non-AIDS (7 AIDS-related complex (ARC); 4 other HIV class IV, 16 asymptomatic) patients. The development of AIDS was preceded 1-4 years by loss or lack of antibody to gag (p15, 24, or 55) and/or to pol (p31, 53, or 64), each p less than 0.001, compared with non-AIDS patients. Correlation between Western blot and recombinant assays was good except in one Western blot p24 (gag) only seroconverter who showed strong reactivity to env by recombinant assay. In conclusion, HIV antibody patterns appear to show prognostic significance in HIV-infected hemophiliacs.
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PMID:Prognostic importance of antibodies to human immunodeficiency virus by recombinant immunoassay and Western blot techniques in HIV antibody-positive hemophiliacs. 316 3

Normal blood donors were examined for human immunodeficiency virus (HIV)-reactive antibodies with both virus- and Escherichia coli-expressed env- and gag-coded antigens. The frequency of samples from normal (low-risk) donors that were repeatedly reactive with an HIV enzyme-linked immunosorbent assay blood screening test (Du Pont Co.) was 0.6%. Two classes of HIV serological reactivity were identified: a minor env-reactive class (0.03 to 0.06% of donors) and the predominant env-nonreactive gag-reactive class (gag reactive only [GRO]) (0.4 to 0.5% of donors). Assignment of env reactivity was made by a synthetic (recombinant) env enzyme-linked immunosorbent assay and virus immunoblot. Most GRO sera reacted with p15/p17 bands on HIV immunoblot. Antibody specificity in GRO sera was confirmed by competition-binding studies with viral gag and E. coli-expressed p55gag. This study provides independent verification that gag-specific antibodies are present in many env-nonreactive sera. More serological and virological studies of individuals with this antibody pattern should be pursued to determine the origin of these gag-reactive antibodies.
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PMID:Antibodies reactive with human immunodeficiency virus gag-coded antigens (gag reactive only) are a major cause of enzyme-linked immunosorbent assay reactivity in a blood donor population. 325 46

Eighty-seven seropositive subjects with HIV (human immunodeficiency virus) infection together with 20 normal controls with no history of any illness were investigated for the presence of circulating immune complexes (CIC) by the conglutinin binding assay (KgBA) and further studied for isotype characterization of CIC. Six out of 87 patients showing very high values for immune complexes (CIC) were studied for the presence of free antigen. In 3 out of 6 (1, IVc1; 1, III; 1, IVa) we could detect by ultracentrifugation analysis the presence of specific HIV (p15) anti-HIV (anti-p15) and gp41-anti-gp41 CIC. Evidence in favour of this finding is supported by: the presence of specific CIC (p15-anti-p15 or gp41-anti-gp41) seen only at pH 7.2; the apparent presence of free antigen and specific HIV antibodies were only at pH 4.0. The relevance of this finding lies in the attempt to explain the occurrence of false seronegativity seen occasionally in symptomatic patients. Thus, the presence of CIC might perhaps interfere in the routine assay (i.e. ELISA) making the diagnosis difficult. All these considerations will have to be taken into account in the future handling of this disease.
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PMID:Characterization of specific immune complexes in HIV-related disorders. 347 75

A current concept of the serological response to human immunodeficiency virus (HIV) infection in humans is that antibodies to core antigens (p55, p24, and p15) are detectable earlier during initial stages of antibody production than antibodies against envelope antigens (gp160, gp120, and gp41). Comparative studies of Western blot (immunoblot), radioimmunoprecipitation assay (RIPA), and enzyme-linked immunosorbent assay (ELISA) during initial antibody production are limited to case reports and have not resolved the issue. Thirty of the 37 participants who are part of a prospective study had at least one specimen that was negative for anti-gp41 but had one or more other bands on Western blot. Twenty-seven of these 30 specimens were reactive for anti-gp120/160 in the RIPA. Of the same 30 specimens, kits from Bionetics identified 2 (7%), ElectroNucleonics 4 (13%), Abbott 13 (43%), Du Pont 25 (83%), and Genetic Systems 25 (83%). All participants had evidence of serological progression by Western blot, including a gp41 band, on subsequent visits; the ELISA kits of all manufacturers identified these later specimens with greater accuracy. These data show that the RIPA detects anti-envelope antibodies that may be not detectable by Western blot and that the production of anti-envelope antibodies approximately parallels the production of anti-core antibodies. The false-negative results by ELISA would permit transmission of HIV by blood transfusion from donors in early stages of infection. The sensitivity of licensed ELISA kits should be improved to identify antibody as soon as possible after infection.
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PMID:Detection of early antibodies in human immunodeficiency virus infection by enzyme-linked immunosorbent assay, Western blot, and radioimmunoprecipitation. 347 69


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