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)

The purpose of this study was to define further the determinants of substrate specificity of HIV-1 PR. Rather than using small peptides, we used an in vitro system which permitted us to evaluate the effect of a mutated site within the context of its natural precursor. We made single-amino-acid substitutions around two sites which are processed by the HIV-1 PR. The Tyr/Pro site within gag appears to encode highly specific determinants which direct proteinase processing between MA and CA. The Phe/Pro site in pol, however, appears to be far more tolerant to amino acid substitutions, as none of our single-amino-acid substitutions blocked cleavage at or around this site. The increased tolerance of the Phe/Pro site may indicate that at this site, structural features are more important determinants of cleavage than primary amino acid sequence. We have shown that sequences outside of those encoding mature PR can inhibit proteolytic processing in this system. By preventing PR from cleaving itself from the polyprotein prematurely, p6* sequences would regulate morphogenesis and infectious particle formation. Late in infection, when the protein concentration of gag and gag/pol polyproteins at the cell surface becomes very high, cooperative protein-protein interactions may cause alterations of a p6*-PR interaction, relieving repression and permitting autocatalysis.
 ...
PMID:Mutational analysis of a native substrate of the HIV-1 proteinase. 166 51

HIV-1 and HIV-2 both cause AIDS in humans. Simian immunodeficiency viruses (SIV) are non-human primate lentiviruses and the closest known relatives of the HIVs. They closely parallel HIVs in genomic organization and biologic properties. The authors discuss the known HIVs and SIVs of African origin and describe the variability which exists in the different groups. HIV-1 and HIV-2 share approximately 55-60% amino-acid homology in gag and pol, the genes most highly conserved among related retroviruses. HIV-1 is spread widely throughout the world, while HIV-2 infection appears to be concentrated in West Africa. Rare cases of HIV-2 infection have, however, been identified in Europe and America, usually in individuals connected with West Africa. The authors discuss viral genetic variation and variation of biological phenotype, and findings on HIV-1 and HIV-2 from Zaire, Uganda, Cameroon, Tanzania, Ethiopia, Congo, Ghana, the Gambia, Guinea-Bissau, and Cote d'Ivoire.
 ...
PMID:Variability among HIV and SIV strains of African origin. 166 22

The gag and pol genes of the human immunodeficiency virus type 1 (HIV-1) (ref. 1) are translated as two polyproteins, Pr55gag and Pr160gag-pol (refs 2-6), which are subsequently cleaved by the action of a virus-encoded protease into the four structural gag proteins of the virion core (p17, p24, p7 and p6) and the pol-encoded enzymes essential for retrovirus replication (protease, reverse transcriptase, ribonuclease H, and endonuclease). Mutational inactivation of the proteases of HIV-1 and other retroviruses results in immature, non-infectious virions, indicating that exogenous inhibition of the protease may represent an attractive approach to anti-AIDS therapy. Here we demonstrate that synthetic peptide analogues, which are potent inhibitors of purified HIV-1 protease, inhibit the processing of the viral polyproteins in cultures of HIV-1-infected T lymphocytes and attenuate viral infectivity.
 ...
PMID:Inhibition of HIV-1 protease in infected T-lymphocytes by synthetic peptide analogues. 168 46

We have modified an Escherichia coli vector expressing 66-kDa HIV-1 reverse transcriptase (p66) so that it simultaneously expresses this and the pol-coded protease. The twin expression cassette yields high quantities of both reverse transcriptase and protease; however, under these conditions, 50% of the over-expressed p66 reverse transcriptase is processed, resulting in accumulation of large quantities of p66/p51 enzyme. Furthermore, addition of a poly(histidine) affinity label at the amino terminus of the reverse-transcriptase-coding sequence (His-p66) permits a simple, rapid purification of milligram quantities of either p66 or p66/p51 enzyme from a crude lysate by metal chelate affinity chromatography. Purified His-p66 and His-p66/His-p51 reverse transcriptase exhibit both reverse transcriptase and RNase H activity. Purification by metal chelate chromatography of a p66/p51 enzyme wherein only the p66 component is labelled strengthens the argument for the existence of a heterodimer.
 ...
PMID:Rapid purification of homodimer and heterodimer HIV-1 reverse transcriptase by metal chelate affinity chromatography. 168 98

We constructed a recombinant vaccinia virus carrying the entire gag and pol genes of human immunodeficiency virus type 1 (HIV-1). The main gene product detected in the lysates of infected CV-1 and SW480 cells was the gag precursor protein. However, in the culture fluid of infected SW480 cells, but not of infected CV-1 cells, reverse transcriptase (RT) activity was detected. The highest RT activity was found at a density of 1.15 g/ml and this fraction contained many round particles with diameters of 100-150 nm. In contrast to the infected cell lysates, the particles contained the processed gag and pol proteins, suggesting that particle formation may be a prerequisite for efficient processing of the gag precursor by the HIV protease encoded in the pol gene. Particles were also recovered from the culture fluid of SW480 cells infected with another recombinant vaccinia virus carrying only the gag gene. These particles contained the unprocessed gag precursor, indicating that the gag precursor alone was sufficient for particle production.
 ...
PMID:Production of human immunodeficiency virus (HIV)-like particles from cells infected with recombinant vaccinia viruses carrying the gag gene of HIV. 168 17

Human immunodeficiency virus type 1 (HIV-1) gag and pol genes were expressed by using fragments of the BH10 clone of HIV inserted into a simian virus 40 late replacement vector. An initial construct containing the entire coding regions of gag, pol, and vif produced only minute amounts of the gag precursor, Pr55gag. However, high-level expression was obtained when an additional sequence from the env gene (the rev-responsive element) was inserted 3' of vif in the correct orientation, and rev was provided in trans from a second vector. Western immunoblot analysis of transfected cells showed the presence of large amounts of both Pr55gag and Pr160gag-pol as well as all of the expected cleavage products. Electron microscopy of thin sections of transfected cells showed a multitude of viruslike particles. Both immature particles in the process of budding and particles containing the condensed core characteristic of HIV were observed. Analysis of the released viruslike particles showed the presence of active reverse transcriptase. Sucrose gradient analysis of particles produced from [3H]uridine-labeled cells indicated a peak of radioactivity which cosedimented with a peak of p24, suggesting that the particles contained RNA.
 ...
PMID:Human immunodeficiency virus type 1 Pr55gag and Pr160gag-pol expressed from a simian virus 40 late replacement vector are efficiently processed and assembled into viruslike particles. 169 47

Two degenerate oligonucleotide primers derived from regions of pol conserved among retroviruses have been synthesized. Polymerase chain reactions utilizing these primers amplify a 135-bp pol fragment in every retrovirus DNA tested to date. The polymerase chain reaction has been linked to a reverse transcriptase step so that a pol-specific DNA fragment can be obtained from a moderate amount of a purified retrovirus or viral RNA. The identity of an unknown retrovirus can be determined by sequencing of the amplified fragment following molecular cloning. This procedure was tested on an unidentified (non-HIV) retrovirus expressed by a B-cell lymphoma line obtained from an AIDS patient. Our PCR assay identified the retrovirus as being highly similar to Mason-Pfizer monkey virus (MPMV) and simian retrovirus 1, which are closely related immunosuppressive type D viruses that cause simian AIDS.
 ...
PMID:The use of primers from highly conserved pol regions to identify uncharacterized retroviruses by the polymerase chain reaction. 169 69

Previous studies revealed that antisense oligodeoxynucleotides to specific regions of the human immunodeficiency virus-1 (HIV-1) are potent inhibitors of replication of HIV-1 in vitro (Zamecnik, P. C., Goodchild, J., Taguchi, Y., and Sarin, P. S. (1986) Proc. Natl. Acad. Sci. U. S. A. 83, 4143-4146). We now report that antisense RNA, synthesized in vitro using T7 and SP6 RNA polymerase, displayed an anti-HIV-1 effect in the HTLV-IIIB/H9 system in vitro. Treatment of HIV-1-infected H9 cells with viral env region antisense RNA encapsulated in liposomes targeted by antibodies specific for the T cell receptor molecule CD3 almost completely inhibited HIV-1 production. The viral env segment covered a part of exon II of HIV-1 tat gene. No anti-HIV activity could be detected with similarly targeted liposome-encapsulated sense env RNA or with pol RNA synthesized in either the sense or antisense orientations, or with env region antisense RNA free in solution, or encapsulated in liposomes in the absence of the targeting antibody. A semiquantitative evaluation revealed that 4000-7000 RNA molecules became cell-bound in targeted liposomes; the half-life of the intracellularly present hybridizable antisense env RNA was approximately 12 h. Western blots showed that antisense env RNA suppressed tat gene expression by approximately 90% and gp160 production by 100%. These data were confirmed by immunoprecipitation studies. Northern blots (using an env probe) demonstrated the existence of all major HIV RNA species (9.3-, 4.3-, and 2.0-kb mRNA) in HIV-infected cells treated with antisense env RNA although at a reduced level. We conclude that the antisense env RNA inhibited viral protein production at the translational level.
 ...
PMID:Inhibition of expression of human immunodeficiency virus-1 in vitro by antibody-targeted liposomes containing antisense RNA to the env region. 169 56

Overexpression of the reverse transcriptase was designed in E. coli. For a high level of expression, HIV protein was expressed as a protein fusion with beta-galactosidase. When the proviral DNA fragment covering the 3' half of the gag gene and the entire pol gene was ligated to the 3' end of the lacZ gene to fuse the truncated gag to lacZ in frame, a small quantity of reverse transcriptase was produced, indicating that frameshifting and post-translational processing have occurred. Much more reverse transcriptase was produced when the entire pol region was directly fused to the lacZ gene. From a one liter culture of bacteria, 1 mg of highly purified reverse transcriptase consisting of approximately equimolar amounts of two species (p64 and p51) was obtained. These proteins had identical N-termini consistent with the deduced amino acid sequence and therefore, might be correctly processed from the fusion protein in E. coli by the protease encoded by the pol region. The purified reverse transcriptase was enzymatically as active as the enzyme purified from the virus particles, and immunoreactive to the sera of HIV carriers with high sensitivity and specificity.
 ...
PMID:Overproduction of human immunodeficiency virus type I reverse transcriptase in Escherichia coli and purification of the enzyme. 169 13

A fragment of the SIVmac251 pol gene was expressed in Escherichia coli as a trpE fusion protein. Analysis of extracts from bacteria containing this expression plasmid revealed the presence of a reverse transcriptase activity dependent on Mg2+ as divalent cation and active on both poly(rA).oligo(dT) and poly(rC.oligo(dG) templates. In comparative studies, the SIV and HIV-1 reverse transcriptases expressed in bacteria displayed very similar high sensitivities to the chain terminator inhibitors AZTTP and ddTTP. The reverse transcriptase of Moloney murine leukemia virus and the DNA polymerase of E. coli were both more resistant to ddTTP, and the E. coli enzyme was significantly more resistant to AZTTP.
 ...
PMID:Expression of enzymatically active reverse transcriptase of simian immunodeficiency virus in bacteria: sensitivity to nucleotide analogue inhibitors. 170 May 44


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