UMLS:C0019693 - FACTA Search

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Query: UMLS:C0019693 (HIV)
170,526 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The binding of p7 nucleocapsid protein of type 1 human immunodeficiency virus (HIV-1) to various oligonucleotides and polynucleotides has been investigated by phosphorescence and optically detected magnetic resonance (ODMR) spectroscopy. The intrinsic spectroscopic probe used in these studies is the photoexcited triplet state of Trp37, which is associated with the C-terminal zinc finger of p7 and is its only tryptophan residue. Complex formation produces a red-shift of the phosphorescence 0, 0-band (DeltaE0,0) of Trp37 as well as a reduction of the zero field splitting (zfs) D parameter. Increases of -DeltaE0,0 (A < C < U < G <I) rank with increasing binding affinity to nucleic acid homooligomers (A approximately C < U < G approximately I). It is proposed that the magnitude of the shift reflects the extent of aromatic stacking interactions. We propose also that -DeltaD increases not only with increased aromatic stacking but also with the extent of charge transfer (CT) character admixed into the triplet state. The quantity DeltaD/DeltaE0,0 correlates with the electron affinity of the bases (G < A < C < U approximately T), suggesting that this quantity reflects the extent of CT character admixed with the triplet state by the aromatic stacking interaction. Also affected by nucleic acid binding of p7 are the kinetic parameters of Trp37. We find a selective increase in the relative populating rate, and of the decay rate constant of the Tx sublevel. In binding of p7 to either d(IT)2 or d(IT)4, two distinct sets of triplet states of Trp37 are resolved, suggesting the existence of specific nucleic acid binding modes of these heterooligomers.
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PMID:Binding of the nucleocapsid protein of type 1 human immunodeficiency virus to nucleic acids studied using phosphorescence and optically detected magnetic resonance. 937 55

The Gag-encoded nucleocapsid protein NCp7 (72 amino acids) from HIV-1, the regulatory protein, Vpr (96 amino acids) and numerous derivatives have been synthesized by solid phase method and their structures determined by 2D NMR. In NCp7, the two highly folded zinc fingers of the Cx2Cx4Hx4C type are in close spacial proximity and the replacement of H by C in the first zinc finger or P by L in the short interdigital domain led to structural modifications evidenced by NMR. In vivo, these point mutations induced a complete loss of viral infectivity by interrupting critical step(s) of the retroviral life cycle. To account for these findings, a model of the complex between NCp7 and d (ACGCC) has been proposed from NMR data, showing the intercalation of Trp37 in the oligonucleotide. This model could also explain the role of NCp7 in the formation of viral particles and agrees with the modifications in morphology of the virions containing mutations in the NCp7 zinc fingers. Vpr is essentially constituted by two long helical domains at its N- and C-terminals and the side chains of Leu60 and Leu67 participate in a leucine-zipper mode of intramolecular interaction. The results obtained have been used to try to develop new antiviral agents inhibiting NCp7 functions and thus possibly devoid of the resistance effects found with inhibitors of HIV enzymes (reverse transcriptase and protease).
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PMID:Structure, biological functions and inhibition of the HIV-1 proteins Vpr and NCp7. 947 50

During reverse transcription of the retroviral genomic RNA, two obligatory DNA strand transfers take place to synthesize the complete proviral DNA with two LTRs. We have previously shown that using an in vitro system made up of two viral RNAs mimicking the 5' and 3' regions of the retroviral genome, both nucleocapsid protein and the repeat (R) sequences were necessary for minus strong-stop cDNA (ss-cDNA) transfer and elongation by reverse transcriptase (RT). In this paper we show that the basic residues of nucleocapsid protein NCp10 of Moloney murine leukemia virus (MoMuLV), but not the zinc finger, are necessary for minus strand transfer. In order to examine the role of the R sequence repeated at the 5' and 3' ends of the genome in minus strand DNA transfer, the MoMuLV R sequence of 68 nt was replaced by either HIV-1 R of 96 nt, or RSV R of 21 nt, or by an artificial sequence of 21 nt. Analysis of MoMuLV DNA strand transfer from the 5' RNA to the 3' RNA and elongation in the presence of NCp10 and RT showed that it was high with control MoMuLV R, high with RSV R, reduced with HIV-1 R, and undetectable with the artificial R sequence. These results suggest that minus strand DNA transfer is a process more complex than simple hybridization of ss-cDNA to the 3' R sequence of the genomic RNA.
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PMID:CIS elements and trans-acting factors required for minus strand DNA transfer during reverse transcription of the genomic RNA of murine leukemia virus. 951 48

The zinc finger structure that is found in the nucleocapsid protein of nearly all retroviruses has been proposed as a target for antiviral therapy. Since compounds that chemically attack the cysteines of the finger have been shown to inactivate both human immunodeficiency virus type 1 (HIV-1) and murine leukemia virus (MuLV) in vitro, 14 of these compounds were tested in an MuLV-induced Friend disease model to assess their ability to inhibit retroviral replication in vivo. Of the 14 compounds tested, only Aldrithiol-2 clearly exhibited anti-retroviral activity as measured indirectly by the delay of Friend disease onset (P < 0.05). These results were confirmed by quantitative competitive polymerase chain reaction studies which monitored viral spread by measuring the level of viral DNA in the peripheral blood mononuclear cells of treated mice. Comparison of treated mice with untreated mice revealed that Aldrithiol-2 produced a greater than 2-log reduction in virus levels. These results functionally demonstrate that a zinc finger-attacking compound can inhibit viral replication in vivo. Since only 1 of the 14 compounds studied was effective, this study also shows the importance of in vivo testing of these types of antiviral compounds in an animal model. Given the strict conservation of the metal-coordinating cysteine structure within HIV-1 and MuLV zinc fingers, our results support the proposal that anti-retroviral drugs which target the nucleocapsid zinc finger may be clinically useful against HIV-1.
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PMID:Inhibition of Friend virus replication by a compound that reacts with the nucleocapsid zinc finger: anti-retroviral effect demonstrated in vivo. 956 28

Affinity capillary electrophoresis was used to detect a shift in mobility when a zinc ion binds to the highly basic nucleocapsid protein (NCp7) of HIV-1. NCp7 contains two Cys-X2- Cys-X4-His-X4-Cys zinc fingers. With constant concentrations of NCp7 as a receptor and various concentrations of zinc as a ligand in the sample buffer and the electrophoresis buffer, we observed changes in electrophoretic mobilities of NCp7 protein when complexes were formed with zinc. Scatchard analysis of the mobility indicates the presence of at least two types of binding sites for zinc. At pH 6.0, one site is shown to bind zinc strongly with a binding constant Kb = 3.25 x 10(5) M-1 and the second site has a Kb = 1.8 x 10(5) M-1. The binding of zinc to the first zinc finger decreased the affinity of zinc for the second zinc finger approximately twofold. The Hill coefficient for this negative cooperativity is 0.9. A series of NCp7 mutants were also examined in the assay to determine their ability to bind zinc. This assay affords a quick method to observe a zinc ion binding to NCp7 and to calculate binding constants.
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PMID:A binding shift assay for the zinc-bound and zinc-free HIV-1 nucleocapsid protein by capillary electrophoresis. 965 80

We injected rat myotubes with proteins and antibodies to assess the importance of the zinc finger (ZnF) domain of the 43-kDa receptor-associated protein, rapsyn, in clustering acetylcholine receptors (AChR). Injection of rat myotubes with a fusion protein containing the ZnF domain of rapsyn disrupted AChR clusters. Clusters were unaffected by a fusion protein containing a double mutant that does not bind zinc. Similar results were obtained with the purified wild type and mutant ZnF domains. The ZnF of HIV-1 nucleocapsid protein had no effect. AChR clusters were also disrupted in myotubes injected with antibodies to the ZnF domain, followed by injection of anti-antibodies. Injection of antibodies directed against a different rapsyn epitope or against the cytoplasmic domain of the AChR had no effect. In transfection experiments with HEK 293 cells, the ZnF domain failed to associate with membrane aggregates containing full-length rapsyn, AChR, or rapsyn and AChR together. We conclude that the ZnF domain of rapsyn provides a binding site essential for AChR clustering, but that this site is unlikely to be involved in high affinity binding of rapsyn to itself or to AChR.
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PMID:The zinc finger domain of the 43-kDa receptor-associated protein, rapsyn: role in acetylcholine receptor clustering. 969 94

The role of the nucleocapsid protein of HIV-1 Gag in virus assembly was investigated using Gag truncation mutants, a nucleocapsid deletion mutant, and point mutations in the nucleocapsid region of Gag, in transfected COS cells, and in stable T-cell lines. Consistent with previous investigations, a truncation containing only the matrix and capsid regions of Gag was unable to assemble efficiently into particles; also, the pelletable material released was lighter than the density of wild-type HIV-1. A deletion mutant lacking p7 nucleocapsid but containing the C-terminal p6 protein was also inefficient in particle release and released lighter particles, while a truncation containing only the first zinc finger of p7 could assemble more efficiently into virions. These results clearly show that p7 is indispensable for virus assembly and release. Some point mutations in the N-terminal basic domain and in the basic linker region between the two zinc fingers, which had been previously shown to have reduced RNA binding in vitro [Schmalzbauer, E., Strack, B., Dannull, J., Guehmann, S., and Moelling, K. (1996). J. Virol. 70: 771-777], were shown to reduce virus assembly dramatically when expressed in full-length viral clones. A fusion protein consisting of matrix and capsid fused to a heterologous viral protein known to have nonspecific RNA binding activity [Ribas, J. C., Fujimura, T., and Wickner, R. B. (1994) J. Biol. Chem. 269: 28420-28428] released pelletable material slightly more efficiently than matrix and capsid alone, and these particles had density higher than matrix and capsid alone. These results demonstrate the essential role of HIV-1 nucleocapsid in the virus assembly process and show that the positively charged N terminus of p7 is critical for this role.
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PMID:The role of nucleocapsid of HIV-1 in virus assembly. 981 10

Although most viral vaccines used in humans have been composed of live attenuated viruses or whole killed viral particles, the latter approach has received little attention in research on experimental primate immunodeficiency virus vaccines. Inactivation procedures involving heat or formalin appear to adversely affect the viral envelope proteins. Recently we have inactivated human immunodeficiency virus type 1 (HIV-1) with the compound 2,2'-dithiodipyridine (Aldrithiol-2, Aldrich, Milwaukee, WI), which inactivates infectivity of retroviruses by covalently modifying the nucleocapsid zinc finger motifs. HIV-1 inactivated with Aldrithiol-2 retained the conformational and functional integrity of the viral and virion-associated cellular proteins on the viral membrane. We have extended our studies of zinc finger targeted inactivation to simian immunodeficiency virus (SIV) and evaluated the feasibility of applying the procedures to large scale (>30 l) production and purification of the primate immunodeficiency viruses. There was no detectable residual infectivity of SIV after treatment with 1 mM Aldrithiol-2 (>5 logs inactivation). Treatment with Aldrithiol-2 resulted in extensive reaction with the nucleocapsid protein of treated virus, as shown by immunoblot and high-performance liquid chromatography (HPLC) analysis. As expected, the virion gp120SU appeared to be completely unreactive with Aldrithiol-2. Sucrose gradient purification and concentration procedures resulted in little loss of viral infectivity or virion-associated gp120SU. When tested in a gp120-CD4 dependent cell binding assay, the inactivated virus bound to cells comparably to the untreated virus. Analysis of gp120-CD4 mediated postbinding fusion events showed that the inactivated virus could induce CD4-dependent fusion with efficiencies similar to the untreated virus. Inactivation and processing of primate immunodeficiency viruses by methods described here results in highly concentrated virus preparations that retain their envelope proteins in a native configuration. These inactivated virus preparations should be useful in whole killed-particle vaccine experiments as well as laboratory reagents to prepare antisera, including monoclonal antibodies, and to study noninfective virion-cell interactions.
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PMID:Chemical inactivation of retroviral infectivity by targeting nucleocapsid protein zinc fingers: a candidate SIV vaccine. 981 59

Retroviral nucleocapsid proteins (NCPs) are CCHC-type zinc finger proteins that mediate virion RNA binding activities associated with retrovirus assembly and genomic RNA encapsidation. Mason-Pfizer monkey virus (MPMV), a type D retrovirus, encodes a 96-amino acid nucleocapsid protein, which contains two Cys-X2-Cys-X4-His-X4-Cys (CCHC) zinc fingers connected by an unusually long 15-amino acid linker. Homonuclear, two-dimensional sensitivity-enhanced 15N-1H, three-dimensional 15N-1H, and triple resonance NMR spectroscopy have been used to determine the solution structure and residue-specific backbone dynamics of the structured core domain of MPMV NCP containing residues 21-80. Structure calculations and spectral density mapping of N-H bond vector mobility reveal that MPMV NCP 21-80 is best described as two independently folded, rotationally uncorrelated globular domains connected by a seven-residue flexible linker consisting of residues 42-48. The N-terminal CCHC zinc finger domain (residues 24-37) appears to adopt a fold like that described previously for HIV-1 NCP; however, residues within this domain and the immediately adjacent linker region (residues 38-41) are characterized by extensive conformational averaging on the micros-ms time scale at 25 degrees C. In contrast to other NCPs, residues 49-77, which includes the C-terminal CCHC zinc-finger (residues 53-66), comprise a well-folded globular domain with the Val49-Pro-Gly-Leu52 sequence and C-terminal tail residues 67-77 characterized by amide proton exchange properties and 15N R1, R2, and (1H-15N) NOE values indistinguishable to residues in the core C-terminal finger. Twelve refined structural models of MPMV NCP residues 49-80 (pairwise backbone RMSD of 0.77 A) reveal that the side chains of the conserved Pro50 and Trp62 are in van der Waals contact with one another. Residues 70-73 in the C-terminal tail adopt a reverse turn-like structure. Ile77 is involved in extensive van der Waals contact with the core finger domain, while the side chains of Ser68 and Asn75 appear to form hydrogen bonds that stabilize the overall fold of this domain. These residues outside of the core finger structure are conserved in D-type and related retroviral NCPs, e.g., MMTV NCP, suggesting that the structure of MPMV NCP may be representative of this subclass of retroviral NCPs.
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PMID:Solution structure and backbone dynamics of Mason-Pfizer monkey virus (MPMV) nucleocapsid protein. 982 93

Nucleocapsid p7 protein (NCp7) zinc finger domains of the human immunodeficiency virus type 1 (HIV-1) are being developed as antiviral targets due to their key roles in viral replication and their mutationally nonpermissive nature. On the basis of our experience with symmetrical disulfide benzamides (DIBAs; Rice et al. Science 1995, 270, 1194-1197), we synthesized and evaluated variants of these dimers, including sets of 4,4'- and 3,3'-disubstituted diphenyl sulfones and their monomeric benzisothiazolone derivatives (BITA). BITAs generally exhibited diminished antiviral potency when compared to their disulfide precursors. Novel, monomeric structures were created by linking haloalkanoyl groups to the benzamide ring through -NH-C(=O)- (amide) or -S-C(=O)- (thiolester) bridges. Amide-linked compounds generally lacked antiviral activity, while haloalkanoyl thiolesters and non-halogen-bearing analogues frequently exhibited acceptable antiviral potency, thus establishing thiolester benzamides per se as a new anti-HIV chemotype. Pyridinioalkanoyl thiolesters (PATEs) exhibited superior anti-HIV-1 activity with minimal cellular toxicity and appreciable water solubility. PATEs were shown to preferentially target the NCp7 Zn finger when tested against other molecular targets, thus identifying thiolester benzamides, and PATEs in particular, as novel NCp7 Zn finger inhibitors for in vivo studies.
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PMID:Synthesis and biological properties of novel pyridinioalkanoyl thiolesters (PATE) as anti-HIV-1 agents that target the viral nucleocapsid protein zinc fingers. 988 34

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