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)

A series of novel distamycin-related polyanionic compounds were compared for their anti-HIV activity. Several were highly potent inhibitors of HIV virus-induced cell killing and viral replication of a wide variety of laboratory isolates, as well as a monocytotropic virus and a clinical isolate in human peripheral blood lymphocytes. These compounds are structurally different from other sulfonic acid containing compounds reported to be potent inhibitors of the human immunodeficiency virus (HIV) in two respects: (1) they are structurally related to the non-toxic minor groove DNA binder distamycin; and (2) a number of them contain the aromatic phosphonic acid group. The compounds that were evaluated can be categorized into monomeric or dimeric ureido structural classes incorporating the bisamido-N-methylpyrrolenaphthalene-sulfonic acid group, with differences in the number and position of the sulfonic acids on the naphthalene rings. Broader structure-activity studies were made possible through the synthesis and evaluation of the compounds containing only a single N-methylpyrrole unit, those incorporating the N-methylpyrazole structure, and compounds having the isosteric phosphonic acid group substituted for the sulfonic acid group. One of the most potent of the inhibitors was 2,2'[4,4'[[aminocarbonyl]amino]bis[N,4'-di[pyrrole-2-carboxamide- 1,1'-dimethyl]]-4,6,8 naphthalenetrisulfonic acid] hexasodium salt, NSC 651015. This compound, the phosphonic acid analog NSC 662162, and the monomeric compound NSC 651018 were studied to determine the mechanism of their inhibitory activity. Mechanistic studies revealed that inhibition was due to the disruption of virus attachment to CD(4+)-susceptible cells and a further restraint on fusion of virus and cell membranes. The relative tolerance of these compounds in mice suggests that sufficient antiviral concentrations could be reached in vivo and thus may prove valuable in the treatment of AIDS patients.
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PMID:Novel sulfonated and phosphonated analogs of distamycin which inhibit the replication of HIV. 854 Jul 54

The nucleocapsid (NC) protein of human immunodeficiency virus type 1 is required for packaging of viral RNA and for virion assembly. It contains two clusters of basic amino acids, consisting of five and four amino acid residues, flanking the first of its two zinc fingers. These amino acid residues have been mutagenized to neutral ones individually, as well as in various combinations, by site-directed mutagenesis. Wild-type NCp7 and the mutant proteins were expressed as recombinant proteins in Escherichia coli, with six histidines as tags at their amino termini in order to allow efficient purification. The purified proteins were analyzed for RNA binding in vitro with human immunodeficiency virus type 1 5' leader RNA transcribed in vitro. Assays comprised Northwestern blots at various salt concentrations and filter binding tests which allowed determination of the dissociation constants of the various mutants. The results indicated that mutations of the amino acid R-7 and of R-32 and K-33 were more critical for RNA binding than other mutations. Mutation of the other amino acid residues reduced the binding affinity in proportion to the number of mutations. Mutation of seven of the nine basic amino acid residues reduced the binding of RNA by 50- to 90-fold.
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PMID:Mutations of basic amino acids of NCp7 of human immunodeficiency virus type 1 affect RNA binding in vitro. 855 14

The activity of human immunodeficiency virus protease is markedly increased at elevated salt concentration. The structural basis of this effect has been explored by several independent methods by using both the wild-type enzyme and its triple mutant (Q7K/L33I/L63I) (Mildner, A. M., Rothrock, D. J., Leone, J. W., Bannow, C. A., Lull, J. M., Reardon, I. M., Sarcich, J. L., Howe, W. J., Tomich, C.-S. C., Smith, C. W., Heinrikson, R. L., and Tomasselli, A. G. (1994) Biochemistry 33, 9405-9413), designed to better resist autolysis. Monitoring the intrinsic fluorescence of the two enzymes during urea-mediated denaturation has shown that at high NaCl concentration, both the conformational stability ( DeltaG0) and the transition midpoint (D1/2) between the folded and unfolded states increase, indicating that the salt stabilizes the enzyme structure. These equilibrium data are supported by kinetic studies on the urea-mediated unfolding by measuring fluorescence change, red shifting in the maximum of the emission spectrum, and far- and near-UV CD. The salt effects observed in urea-mediated unfolding reactions prevail upon heat denaturation. All these findings support the existence of a two-state equilibrium between the folded and unfolded proteins. The pH dependence of fluorescence intensity indicated that the conformation of human immunodeficiency virus type 1 protease should change in the catalytically competent pH region. It is concluded that preferential hydration stabilizes the protease structure in the presence of salt, providing entropic contribution to enhance the catalytic activity.
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PMID:Conformational stability and catalytic activity of HIV-1 protease are both enhanced at high salt concentration. 862 2

Continuous replication of human immunodeficiency virus type 1 requires the expression of the regulatory protein Rev, which binds to the Rev response element (RRE) and up-regulates the cytoplasmic appearance of singly spliced and unspliced mRNA species. It has been demonstrated that the murine protein YL2 interacts with Rev in vivo and modulates the activity of Rev (Luo, Y., Yu, H., and Peterlin, B. M. (1994) J. Virol. 68, 3850-3856). Here we show that the YL2 human homologue, the p32 protein, which co-purifies with alternative splicing factor ASF/SF2, interacts directly with the basic domain of Rev in vitro and that the Rev-p32 complex is resistant to high concentrations of salt or nonionic detergent. Protein footprinting data suggest that Rev interacts specifically with amino acids within the 196-208 region of p32. An analysis of the ternary complex, formed among p32, Rev, and RRE RNA, shows that Rev can bridge the association of p32 and RRE. Furthermore, we demonstrate that exogenously added p32 specifically relieves the inhibition of splicing in vitro exerted by the basic domain of Rev. Our data are consistent with a model in which p32 functions as a link between Rev and the cellular splicing apparatus.
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PMID:In vitro interaction between human immunodeficiency virus type 1 Rev protein and splicing factor ASF/SF2-associated protein, p32. 862 63

The three-dimensional structure of the amino-terminal core domain (residues 1 through 151) of the human immunodeficiency virus-type 1 (HIV-1) capsid protein has been solved by multidimensional heteronuclear magnetic resonance spectroscopy. The structure is unlike those of previously characterized viral coat proteins and is composed of seven alpha helices, two beta hairpins, and an exposed partially ordered loop. The domain is shaped like an arrowhead, with the beta hairpins and loop exposed at the trailing edge and the carboxyl-terminal helix projecting from the tip. The proline residue Pro1 forms a salt bridge with a conserved, buried aspartate residue (Asp51), which suggests that the amino terminus of the protein rearranges upon proteolytic maturation. The binding site for cyclophilin A, a cellular rotamase that is packaged into the HIV-1 virion, is located on the exposed loop and encompasses the essential proline residue Pro90. In the free monomeric domain, Pro90 adopts kinetically trapped cis and trans conformations, raising the possibility that cyclophilin A catalyzes interconversion of the cis- and trans-Pro90 loop structures.
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PMID:Structure of the amino-terminal core domain of the HIV-1 capsid protein. 866 5

The construction, expression, and purification of an active Fv fragment of the 0.5beta monoclonal human immunodeficiency virus type 1 (HIV-1) neutralizing antibody is reported. The interaction between the Fv fragment and the RP135 peptide derived from the V3 loop of gp120 from HIV-1IIIB was studied by varying the salt concentration and by mutating arginine residues in the peptide. The mutations R4A, R8A and R11A (which correspond to residues 311, 315, and 318 in gp120 of HIV-1IIIB) reduce the binding free energy by 0.22 (+/- 0. 20), 4.32 (+/- 0.16), and 1.58 (+/- 0.17) kcal mol-1, respectively. The salt-dependent components of their contributions to binding are 0.02 (+/- 0.22), -0.55 (+/- 0.18), and -0.97 (+/- 0.19) kcal mol-1, respectively. The magnitudes of the mutational effects and the extent of shielding by 1 M NaCl suggest that Arg-8 is involved in a buried salt bridge in the peptide-Fv fragment complex, whereas Arg-11 is involved in a more solvent-exposed electrostatic interaction.
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PMID:Contribution of arginine residues in the RP135 peptide derived from the V3 loop of gp120 to its interaction with the Fv fragment of the 0.5beta HIV-1 neutralizing antibody. 866 80

A biologically contained cytoprotection assay was developed to screen inhibitors of the human immunodeficiency virus without the need for high level containment or practices. The virus used has multiple point mutations that have destroyed its ability to produce both Rev and Tat, proteins essential for virus replication in vitro. The original cell line employed (CEM-SSTART) contains a genetic construct that allows for the continuous expression of both Rev and Tat, and a subclone (1A2) was developed that provides for maximum acute cytopathic effect. The National Cancer Institute's AIDS drug screening assay was used to test known drugs with both HIVIIIB virus in the T4 lymphocytic cell line CEM-SS and mutant virus in the 1A2 subclone. This cell-based assay uses the tetrazolium salt, XTT, as an indicator of cellular metabolism after the cells have been infected with virus. The results of extensive testing have shown that the assay using mutant virus is comparable to the current NCI AIDS drug screen. After 42 days in 1A2 or CEM-SS cell culture, the virus or the integrated genome did not revert to wild-type, and the virus produced in 1A2 cells was unable to replicate in PBMCs. Mutant viral stocks were devoid of wild-type virus as determined by a PCR assay that would have found 60-600 copies of mutant RNA. These materials, which are now available to the scientific community (NIH AIDS Research and Reference Reagent Program), should be useful tools to screen and test compounds for potential inhibition of HIV in laboratories not equipped to maintain and use wild-type infectious virus.
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PMID:Assessment of a cytoprotection assay for the discovery and evaluation of anti-human immunodeficiency virus compounds utilizing a genetically-impaired virus. 878 55

The human immunodeficiency virus type-1 (HIV-1) encodes a protease which is essential for the production of infectious virus. The protease prefers substrates that contain glutamic acid or glutamine at the P2' position. The catalytic role of these residues has been studied by using a highly specific fluorogen substrate, 2-aminobenzoyl-Thr-Ile-Nle-Phe(NO2)-Gln-Arg (substrate QR), and its counterpart (substrate ER) containing Glu in place of Gln. The newly designed substrate ER that contains a pair of charged residues at P2' and P3' sites is the most specific substrate described so far for HIV-1 protease. The specificity rate constant (kcat/Km = 2.1 x 10(7) M-1 s-1) approaches, but does not reach, the diffusion limit. This follows from the appreciable solvent kinetic deuterium isotope effects on the rate constants, indicating that, independent of the salt concentration, the rate-limiting step of the catalysis is a chemical process rather than a physical one. The reaction also has positive entropy of activation. On the other hand, the rate-limiting step for substrate QR changes with increasing salt concentration from a physical to chemical step, while the negative activation entropy becomes positive. The rate increase with substrate ER is 50-fold with respect to substrate QR in the presence of 0.1 M NaCl and diminishes to 3.5-fold at 2.0 M NaCl concentration, as a consequence of a considerable rate increase at high salt concentration with substrate QR but not with substrate ER. The Km value is much lower for the substrate ER (0.8 microM) than for substrate QR (15 microM), indicating a more effective binding for substrate ER at 0.1 M NaCl. Unexpectedly, the strong binding appears to be achieved by the unionized form of Glu in P2', as follows from the remarkably different pH-rate profiles for substrates QR and ER. The effective binding elicited by the glutamic acid may be utilized in designing inhibitors for therapeutic purposes.
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PMID:Rate-determining steps in HIV-1 protease catalysis. The hydrolysis of the most specific substrate. 894 73

The tethered-dimer protease of human immunodeficiency virus 1 (HIV-1) [Cheng Y.-S. E., Yin, F.H., Foundling, S., Blomstrom, D. & Kettner, C. A. (1990) Proc. Natl Acad. Sci. USA 87, 9660-9664] and its mutants containing amino acid substitutions or deletions or both in only one flap region were expressed in Escherichia coli. These mutant enzymes showed various degrees of self-processing and significantly reduced catalytic activity toward oligopeptide substrates compared with the wild type. Kinetic parameters determined for one of the oligopeptide substrates showed a dramatic increase in K(m) and decrease in Kcat values. Unexpectedly, the substrate cleavage was more efficient in low salt concentration for a mutant containing a shortened hydrophilic flap. Assays with oligopeptides representing naturally occurring cleavage sites or oligopeptides containing single amino acid substitutions at the P2 and P2' substrate positions showed only moderate changes in the substrate specificity of the mutant proteases. Predicted structures for the mutants were constructed by molecular modeling and used to interpret the results of kinetic measurements. In general, the data suggest that the mutated part of the flaps does not have a major role in determining substrate specificity; rather, it provides the hydrophobic environment and hydrogen-bond interactions with the conserved water that are necessary for efficient substrate binding and catalysis.
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PMID:Activity of tethered human immunodeficiency virus 1 protease containing mutations in the flap region of one subunit. 906 69

Host site selection for full-site integration by human immunodeficiency virus type-1 (HIV-1) intergrase (IN) from nonionic detergent-lysed virions was investigated. Linear retrovirus-like DNA (469 bp) possessing 3' OH recessed long terminal repeat termini was efficiently inserted by a bimolecular donor reaction into a supercoiled DNA target (2867 bp), producing the HIV-1 5-bp host site duplication. Sequence data were analyzed from 193 donor-target recombinants obtained from the linear 3.8-kb DNA product. The selection of host target sites appeared randomly distributed and was independent of lysis and assay conditions. The fidelity of the 5-bp duplications in comparison to other size duplications was highest (94%) with high-salt (300 mM NaCl) lysis of the virions and 60 mM NaCl for strand transfer using Mg2+ as the divalent cation. Base sequence analysis demonstrated some biases in the 5-bp duplications at the sites of strand transfer and at the immediate host sequences surrounding the duplications. In addition to the observed duplications, approximately 30% of the recombinants isolated from the linear 3.8-kb DNA product contained specific and repetitive small-size deletions. No deletions smaller that 17 bp were observed and the distance between the deletion sets had a periodicity of approximately 10 bp. The mechanisms involved in how HIV-1 IN produces the 5-bp duplications and the repetitive host site deletions are discussed.
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PMID:Host site selection for concerted integration by human immunodeficiency virus type-1 virions in vitro. 916 83


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