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

The relatively fast artificial substrate Leu-Ser-rho-nitro-Phe-Nle-Ala-Leu-OMe generates a solvent isotope effect of 1.51 +/- 0.02 only on the maximal velocity of peptide hydrolysis catalyzed by porcine pepsin (EC 3.4.23.1). The absence of an isotope effect on V/K places the isotopically-sensitive step after peptide bond cleavage and the release of the first product. Reprotonation of the active site aspartic carboxyls is proposed as the most likely interpretation of this observation. Structural and kinetic similarities between pepsin and other aspartic proteinases, including the therapeutically important targets HIV protease and renin, suggest a similar slow reprotonation step after catalysis. This mechanistic feature has important implications regarding inhibitor design; if most of the enzymes are present in a product-release form during steady-state turnover, then perhaps inhibitors should be designed as product analogs instead of substrate analogs.
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PMID:Slow step after bond-breaking by porcine pepsin identified using solvent deuterium isotope effects. 201 46

Mutations were introduced into the P2 and P1 positions of the junctions, (a) linking reverse transcriptase (RT) and integrase (IN) (-Leu*Phe-) and (b) between the p51 and RNase H domain (-Phe*Tyr-) within p66 of RT in the HIV-1 pol polyprotein. Processing by HIV proteinase (PR) in cis was monitored upon expression of these constructs in E. coli. Whereas the presence of Leu or Phe in P1 permitted rapid cleavage at either junction, substitution of a beta-branched (Ile) hydrophobic residue essentially abolished hydrolysis. By contrast, placement of a beta-branched (Val) residue in the P2 position flanking such -Hydrophobic*Hydrophobic- junctions resulted in effective cleavage of the scissile peptide bond. Gly in P2, however, abrogated cleavage. The significance of these findings in terms of PR specificity, polyprotein processing and the generation of homodimeric (p51/p51) RT for crystallisation purposes is discussed.
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PMID:Mutating P2 and P1 residues at cleavage junctions in the HIV-1 pol polyprotein. Effects on hydrolysis by HIV-1 proteinase. 204 56

Novel fluorogenic substrates for human immunodeficiency viral protease have been developed based on the principle of fluorescence energy transfer. Starting from a p24/p15 cleavage site-derived hexapeptide substrate. Ac-Thr-Ile-Nle-Nle-Gln-Arg-NH2, incorporation of 2-aminobenzoic acid in place of the acetyl group as the donor and p-NO2-Phe at the P1' position as acceptor gave the intramolecularly quenched fluorogenic substrate. Cleavage of the substrate by HIV protease released the fluorescent N-terminal tripeptide from its close apposition to the quenching nitrobenzyl group, resulting in enhanced fluorescence. An automated assay based on 96-well microtiter plates and a fluorometric plate reader have been developed, which allow high throughput of compounds in the search for HIV protease inhibitors.
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PMID:A simple, continuous fluorometric assay for HIV protease. 209 Jun 47

A new method is described for determining molecular structures from NMR data. The approach utilizes 2D NOESY back-calculations to generate simulated spectra for structures obtained from distance geometry (DG) computations. Comparison of experimental and back-calculated spectra, including analysis of cross-peak buildup and auto-peak decay with increasing mixing time, provides a quantitative measure of the consistence between the experimental data and generated structures and allows for use of tighter interproton distance constraints. For the first time, the "goodness" of the generated structures is evaluated on the basis of their consistence with the actual experimental data rather than on the basis of consistence with other generated structures. This method is applied to the structure determination of an 18-residue peptide with an amino acid sequence comprising the first zinc fingerlike domain from the gag protein p55 of HIV. This is the first structure determination to atomic resolution for a retroviral zinc fingerlike complex. The peptide [Zn(p55F1)] exhibits a novel folding pattern that includes type I and type II NH-S tight turns and is stabilized both by coordination of the three Cys and one His residues to zinc and by extensive internal hydrogen bonding. The backbone folding is significantly different from that of a "classical" DNA-binding zinc finger. Residues C(1)-F(2)-N(3)-C(4)-G(5)-K(6) fold in a manner virtually identical with the folding observed by X-ray crystallography for related residues in the iron domain of rubredoxin; superposition of all main-chain and Cys side-chain atoms of residues C(1)-K(6) of Zn(p55F1) onto residues C(6)-Y(11) and C(39)-V(44) of rubredoxin gives RMSDs of 0.46 and 0.35 A, respectively. The side chains of conservatively substituted Phe and Ile residues implicated in genomic RNA recognition form a hydrophobic patch on the peptide surface.
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PMID:High-resolution structure of an HIV zinc fingerlike domain via a new NMR-based distance geometry approach. 210 40

Because polymorphonuclear neutrophils are the most important component of host defense against bacteria, we assessed their function in 13 children with asymptomatic and 12 with symptomatic infection with human immunodeficiency virus type 1 (HIV-1), and compared their values with healthy adult control values. The functions assessed were (1) chemotaxis, (2) bacterial phagocytosis, (3) superoxide generation, and (4) bactericidal activity. Chemotaxis of polymorphonuclear neutrophils toward the chemoattractant N-formylmethionyl leucyl phenylalanine (FMLP) was significantly decreased in symptom-free infected children compared with control subjects (p less than 0.0001), but was increased in children with symptomatic infection (p less than 0.025). Bactericidal activity of the neutrophils against Staphylococcus aureus was defective in 8 of 12 children with asymptomatic infection (p = 0.016), and in 8 of 9 children with symptomatic infection (p less than 0.00001). Superoxide generation by polymorphonuclear neutrophils on stimulation with FMLP and phagocytosis of S. aureus were normal. Serum from patients with symptomatic HIV-1 infection was not as efficient in low concentrations as normal serum in the ability to opsonize S. aureus. The in vitro bactericidal defect was partially corrected by granulocyte-macrophage colony-stimulating factor (GM-CSF). The results suggest that both cellular (neutrophils) and humoral defects contribute to the increased incidence of bacterial infections in HIV-1-infected children, and that GM-CSF may improve the defective bactericidal activity of polymorphonuclear neutrophils in these patients.
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PMID:Impairment of neutrophil chemotactic and bactericidal function in children infected with human immunodeficiency virus type 1 and partial reversal after in vitro exposure to granulocyte-macrophage colony-stimulating factor. 217 Jun 9

Dideoxynucleosides (zidovudine[AZT], dideoxycytidine[ddC], and dideoxyinosine[ddI]) are promising new agents for the management of human immunodeficiency virus type 1 (HIV-1) infections. In light of recent data demonstrating defects in the polymorphonuclear leukocyte (PMN) bactericidal activity of HIV-1-infected patients and since many chemotherapeutic agents affect PMN function, we examined their effects on the function of PMNs from both healthy and HIV-1-infected individuals in vitro. AZT (0.1 to 25 microM), ddC (0.01 to 1 microM), and ddI (0.2 to 50 microM) had no effect on viability, chemotaxis to N-fromylmethionyl leucyl phenylalanine, phagocytosis of Candida albicans or Staphylococcus aureus, or superoxide production following stimulation by N-formylmethionyl leucyl phenylalanine. Killing of C. albicans was not affected by AZT but was enhanced by 0.1 and 1 microM ddc (a 1 microM, killing was 26.0 +/- 2.02% compared with 17.0 +/- 0.73% for controls: P = 0.006) and 0.2 to 50 microM ddI (at 10 microM, killing was 25.0 +/- 0.68% compared with 17.8 +/- 0.91% for controls; P = 0.002). Killing of S. aureus was unchanged by AZT and ddC but was significantly enhanced by ddI at 0.2 to 20 microM (at 2 microM, killing was 71.2 +/- 5.57% compared with 51.4 +/- 6.29% for controls; P = 0.0045). In addition, the preexisting defective bactericidal capacity of PMNs from HIV-1-infected patients was enhanced by ddI (P less than 0.025). Potential enhancement by these dideoxynucleosides of certain PMN functions of HIV-1-infected patients deserves further study.
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PMID:Effects of antiretroviral dideoxynucleosides on polymorphonuclear leukocyte function. 217 34

The aspartyl protease of human immunodeficiency virus 1 (HIV-1) has been expressed in Escherichia coli at high levels, resulting in the formation of inclusion bodies which contain denatured insoluble aggregates of the protease. After solubilization of these inclusion bodies in guanidinium chloride, the protease was purified to apparent homogeneity by a single-step reverse-phase HPLC procedure. The purified, but inactive, protein was denatured in 8 M urea and refolded to produce the active protease. Enzyme activity was demonstrated against the substrate H-Val-Ser-Gln-Asn-Tyr-Pro-Ile-Val-OH, modeled after the cleavage region between residues 128 and 135 in the HIV gag polyprotein. With this substrate, a Vmax of 1.3 +/- 0.2 mumol/(min.mg) and KM of 2.0 +/- 0.3 mM were determined at pH 5.5. Pepstatin (Iva-Val-Val-Sta-Ala-Sta-OH) and substrate analogues with the Tyr-Pro residues substituted by Sta, by Phe psi [CH2N]Pro, and by Leu psi [CH(OH)CH2]Val inhibited the protease with KI values of 360 nM, 3690 nM, 3520 nM, and less than 10 nM, respectively. All were competitive inhibitors, and the tightest binding compound provided an active site titrant for the quantitative determination of enzymatically active HIV-1 protease.
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PMID:Substrate analogue inhibition and active site titration of purified recombinant HIV-1 protease. 218 16

By replacement of the P1' residue in a capsid/nucleocapsid cleavage site mimic with 4-NO2-phenylalanine (Nph), an excellent chromogenic substrate, Lys-Ala-Arg-Val-Leu*Nph-Glu-Ala-Met, for HIV-1 proteinase (kappa cat = 20 s-1, Km = 22 microM) has been prepared. Substitution of the Leu residue in P1 with norleucine, Met, Phe, or Tyr had minimal effects on the kinetic parameters (kappa cat and kappa cat/Km) determined at different pH values, whereas peptides containing Ile or Val in P1 were hydrolyzed extremely slowly. The spectrophotometric assay has been used to characterize the proteinase further with respect to pH dependence, ionic strength dependence, and the effect of competitive inhibitors of various types.
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PMID:Sensitive, soluble chromogenic substrates for HIV-1 proteinase. 218 27

We report for the first time the concentrations of free amino acids in human intestinal biopsies obtained by routinely performed endoscopy. We studied 15 medical patients with no changes of the mucosa and six HIV-infected persons with duodenitis. The mean (and SD) sum of all amino acids, taurine excepted, was 61.9 (5.4) mmol/kg dry weight in duodenal biopsies of HIV-negative subjects (n = 11) and 82.9 (0.6) mmol/kg in colonic specimens: 50% (44%) of the total (minus taurine) consisted of aspartate and glutamate and 14% (12%), of the essential amino acids. The relative amino acid pattern in duodenum and colon differed completely from that for muscle: aspartate was fourfold higher; glutamate, phenylalanine, glycine, valine, leucine, and isoleucine were about twofold higher. In contrast, glutamine amounted only to 4% (duodenum) to 14% (colon) of muscle glutamine. In duodenal biopsies of the HIV-infected persons, we found significantly (P less than 0.01, except glutamine: P less than 0.025) increased concentrations of glutamate (24.1 vs 17 mmol/kg dry weight), ornithine (1.4 vs 0.4), valine (2.2 vs 1.7), and glutamine.
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PMID:Intracellular free amino acid patterns in duodenal and colonic mucosa. 230 84

Ac-Lys-Ala-Ser-Gln-Asn-Phe(NO2)-Pro-Val-Val-NH2 (peptide I) and Thr-Phe-Gln-Ala-Phe(NO2)-Pro-Leu-Arg-Glu-Ala (peptide II) undergo hydrolysis between the p-nitrophenylalanyl and prolyl residues catalyzed by the proteases of HIV-1 and AMV, respectively. The specific hydrolyses of peptides I and II are accompanied by a decrease in their uv absorption at 269 nm (delta epsilon = 1000) and an increase at 316 nm (delta epsilon = 600). The use of microspectrophotometric cells allows continuous uv measurements on a volume (60 to 120 microliters) comparable to that required for the HPLC point assay currently used. At the highest substrate concentration possible under the assay conditions, good first-order kinetics were observed with both proteases, and the values of Vmax/Km were obtained.
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PMID:Continuous spectrophotometric assay for retroviral proteases of HIV-1 and AMV. 255 Dec 68


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