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)

The nef genes, derived from two different human immunodeficiency-virus-type-1 (HIV-1) strains, were expressed in procaryotic cells (Escherichia coli) and in eucaryotic cells (insect cells infected with nef-containing baculovirus). The oligomerization of recombinant Nef protein was studied by NMR spectroscopy and immunoblotting under various experimental conditions. 1H-NMR spectroscopy shows that native folded protein has the tendency to polymerize under low-salt conditions. These oligomers become covalently linked by disulfide bonds after decreasing the reduction potential, a process which is fully reversible. Cross-linking studies with bis(sulfo-succinimidyl)suberate and alkylation with iodoacetic acid under non-reducing and reducing conditions document for the first time that Nef can also form homomeric structures including monomers, dimers, trimers and tetramers in cell lysates and intact cells. We found disulfide-linked as well as non-covalently associated oligomers. Since the Nef molecules are not exclusively found in the cytoplasm of HIV infected cells and since the reduced glutathione concentration in lymphocytes of virus infected persons is known to be unusually low, it might be possible that these Nef oligomers have a biological function in vivo as well.
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PMID:Oligomerization of the Nef protein from human immunodeficiency virus (HIV) type 1. 851 95

The Rev Response Element (RRE) RNA-Rev protein interaction is important for regulation of gene expression in the human immunodeficiency virus. A model system for this interaction, which includes stem IIB of the RRE RNA and an arginine-rich peptide from the RNA-binding domain of Rev, was studied using multidimensional heteronuclear NMR. Assignment of the RNA when bound to the peptide was obtained from NMR experiments utilizing uniformly and specifically 13C-labeled RNA. Isotopic filtering experiments on the specifically labeled RNA enable unambiguous assignment of unusual nonsequential NOE patterns present in the internal loop of the RRE. A three-dimensional model of the RNA in the complex was obtained using restrained molecular dynamics calculations. The internal loop contains two purine-purine base pairs, which are stacked to form one continuous helix flanked by two A-form regions. The formation of a G-G base pair in the internal loop requires an unusual structure of the phosphate backbone. This structural feature is consistent with mutational data as being important for the binding of Rev to the RRE. The G-G base pair may play an important role in opening the normally narrow major groove of A-form RNA to permit binding of the Rev basic domain.
J Biomol NMR 1995 Dec
PMID:Assignment and modeling of the Rev Response Element RNA bound to a Rev peptide using 13C-heteronuclear NMR. 856 66

Synthetic peptide antigens corresponding to the entire third variable region V3, the principal neutralizing determinant of the human immunodeficiency virus (HIV) envelope glycoprotein of HIV-1 subtype B (1), HIV-2 subtype A (5), and HIV-2 subtype B (7) were synthesized by solid-phase peptide synthesis (Table 1). 1 and 5 were also prepared as their GlcNAc-glycosylated forms at the natural N-glycosylation site NXT (positions 6-8; peptides 4 and 6). Additionally, the proposed beta-turn region of 1 (GPGR; positions 15-18) was altered by introducing D-Ala17 (2) and D-Pro16 (3). All compounds have been studied by two-dimensional NMR techniques. Interproton distances and 3JNH/H alpha coupling constants derived from NMR data are used as restraints in distance geometry and ENSEMBLE-Distance and angle-bound driven dynamics calculations. The stimulations led to disordered conformations except for a high propensity of a beta II-turn in the region GPXR (positions 15-18) in 1, 2, and 4. In 3 (G-D-ProGR, positions 15-18), a type beta I'-turn was mainly found instead. For peptide 7, the consensus sequence of HIV-2 subtype B, a type beta II-turn was also found although the primary structure (VSGL; positions 15-18) differs grossly from the HIV-1 peptide 1. With the exception of 2, all beta II-turns were able to form a canonically opened beta-turn by a 180 degree rotation of phi(G17). Surprisingly, compounds 5 and 6 that are highly similar to 7 showed no beta II-type turn within MSGL (positions 15-18). They form a type beta VIII-turn across the tetrapeptide SGLV (positions 16-19) together with a non-canonical turn conformation across LMSG (positions 14-17) leading to an S-conformation. The reaction of the peptides with HIV-positive sera from patients infected with different subtypes of HIV-1 and HIV-2 was tested in enzyme-linked immunosorbent assays (ELISA reactions). No HIV-2 sera reacted peptide 1 and no HIV-1 sera showed reactivity to peptide 5. We propose that certain amino acid exchanges within the V3 domain lead to altered conformations of the V3 loop resulting in antibodies that show altered binding properties to the peptide antigens used in the ELISA reactions.
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PMID:Secondary structural elements as a basis for antibody recognition in the immunodominant region of human immunodeficiency viruses 1 and 2. 862 Aug 73

The frameshift protein p6* encoded directly upstream of the protease in the human immunodeficiency virus type 1 (HIV-1) pol reading frame is thought to be a natural inhibitor of protease activation and to play a role in the polyprotein processing of Gag and Gag-Pol precursors. To allow structural characterization of the p6* transframe protein, the p6* coding region was cloned into the vector pGEX-KG and expressed in Escherichia coli as a fusion protein with glutathione S-transferase (GST) under the control of the tac promoter. Thrombin cleavage of the construct resulted in a 70-amino-acid polypeptide which is extended by two additional residues at the N-terminus compared to the natural p6* sequence. The native purification procedure including an affinity and a size-exclusion chromatography step yielded sufficient amounts of highly pure protein suitable for NMR spectroscopy. Fluorescence, circular dichroism and 1H-NMR spectroscopy were applied to characterize the structure of protein. Two-dimensional NMR spectra provided essentially complete sequence-specific resonance assignments at pH 5.9. Although there is evidence for a helix-forming tendency in the N-terminus of the protein, the experiments indicate that p6* has no overall stable secondary or tertiary structure with the single tryptophan exposed in aqueous solution. However, the results reported herein open the way to characterize further the interaction of p6* with the HIV-1 protease in structural and functional in vitro studies.
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PMID:Sequence-specific resonance assignments of the 1H-NMR spectra and structural characterization in solution of the HIV-1 transframe protein p6. 864 76

Specific binding of the human immunodeficiency virus Tat protein to its RNA site (TAR) is mediated largely by a single arginine residue located within a basic region of the protein. Many essential features of the interaction can be mimicked by the free amino acid arginine, and an NMR model has been proposed in which the arginine guanidinium group binds to a guanine base in the major groove and to two phosphates adjacent to a bulge, with the RNA structure stabilized by a base triple between a U in the bulge and an adjacent A:U base pair. To compare the TAR structure to other arginine-binding RNAs, we performed in vitro selection experiments and identified RNAs with arginine-binding affinities similar to TAR. About 40% of the selected RNAs contained the same motif found in TAR: two stems separated by a bulge of at least two nucleotides, a U at the 5' position of the bulge, and G:C and A:U base pairs above the bulge. In many cases, the upper stems contained only the G:C and A:U pairs, located next to small loops. Chemical modification experiments demonstrated that these "TAR-like" RNAs bound arginine in a manner similar to TAR, and in some cases identified nucleotides outside the binding site that contributed to binding. To explore how small loops might help stabilize the structures of adjacent arginine-binding sites, we measured arginine-binding affinities of TAR-like RNAs having all possible three-nucleotide loops. An RNA with a UAG loop bound with highest affinity, and chemical modification and RNase mapping experiments suggested that the RNA changes conformation upon arginine binding, converting a large unstructured loop into a bulge conformation related to that of TAR. The results suggest that the arginine-binding site in TAR is structurally versatile and demonstrate how binding can be modulated by the surrounding RNA context.
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PMID:Arginine-binding RNAs resembling TAR identified by in vitro selection. 865 64

The structures of four different RNA pseudoknots that provide one of the signals required for ribosomal frameshifting in mouse mammary tumor virus have been determined by NMR. The RNA pseudoknots have similar sequences and assume similar secondary structures, but show significantly different frameshifting efficiencies. The three-dimensional structures of one frameshifting and one non-frameshifting RNA pseudoknot had been determined previously by our group. Here we determine the structures of two new RNA pseudoknots, and relate the structures of all four pseudoknots to their frameshifting abilities. The two efficient frameshifting pseudoknots adopt characteristic bent conformations with stem 1 bending towards the major groove of stem 2. In contrast, the two poor frameshifting pseudoknots have structures very different from each other and from the efficient frameshifters. One has linear, coaxially stacked stems, the other has stems twisted and bent, but in the opposite direction to the efficient frameshifters. Changes in loop size that favor bending (shorter loops) increase frameshifting efficiency; longer loops that allow linear arrangement of the stems decrease frameshifting. Frameshifting pseudoknots in feline immunodeficiency virus and simian retrovirus have different loop sequences, but the sequences at their stem junctions imply the same bent conformation as in the mouse mammary tumor viral RNA. The requirement for a precise pseudoknot conformation for efficient frameshifting strongly implies that a specific interaction occurs between the viral RNA pseudoknot and the host protein-synthesizing machinery.
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PMID:A characteristic bent conformation of RNA pseudoknots promotes -1 frameshifting during translation of retroviral RNA. 875 14

TAT protein is an essential regulatory protein of the human immunodeficiency virus type 1 (HIV-1). Inhibition of TAT activity blocks the virus cycle, and a drug that blocks TAT is one of the possibilities to cure AIDS. Circular dichroism (CD) was measured for TAT peptides covering the TAT sequence with overlaps. The CD spectrum of each peptide was measured in different solvents to evaluate the ability of each TAT region to form different secondary structures. The most variation or conformational heterogeneity is observed with the two regions adjacent to the TAT basic region. CD data show that the basic region can adopt an extended structure in a full TAT protein, which is not the case for the isolated peptide. TAT sequences from the different HIV-1 isolates were analyzed, and the results showed that the sequences could be gathered into six groups. Molecular modeling was done on the various isolates based on a TAT structure from two-dimensional NMR. After minimization and dynamic steps, the modeled three-dimensional structures were compared. The results showed structural variations of the TAT protein as a function of the HIV-1 isolates. These structural variations were mainly in the two regions adjacent to the basic region, confirming the conformational heterogeneity indicated by the CD measurements. Furthermore, Chou-Fasman analysis shows significant changes in propensities for each secondary structure only for regions III and V. This conformational heterogeneity should be essential for TAT activity and points out that regions III and V are a poor potential target to design a TAT ligand. We propose a target involving TAT structurally conserved regions, accessible whatever the size of the TAT C terminus.
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PMID:Conformational heterogeneity in two regions of TAT results in structural variations of this protein as a function of HIV-1 isolates. 879 35

Efficient transcription from the human immunodeficiency virus (HIV) promoter depends on binding of the viral regulatory protein Tat to a cis-acting RNA regulatory element, TAR. Tat binds at a trinucleotide bulge located near the apex of the TAR stem-loop structure. An essential feature of Tat-TAR interaction is that the protein induces a conformational change in TAR that repositions the functional groups on the bases and the phosphate backbone that are critical for specific intermolecular recognition of TAR RNA. We have previously determined a high resolution structure for the bound form of TAR RNA using heteronuclear NMR. Here, we describe a high resolution structure of the free TAR RNA based on 871 experimentally determined restraints. In the free TAR RNA, bulged residues U23 and C24 are stacked within the helix, while U25 is looped out. This creates a major distortion of the phosphate backbone between C24 and G26. In contrast, in the bound TAR RNA, each of the three residues from the bulge are looped out of the helix and U23 is drawn into proximity with G26 through contacts with an arginine residue that is inserted between the two bases. Thus, TAR RNA undergoes a transition from a structure with an open and accessible major groove to a much more tightly packed structure that is folded around basic side chains emanating from the Tat protein.
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PMID:Structure of HIV-1 TAR RNA in the absence of ligands reveals a novel conformation of the trinucleotide bulge. 891

The three-dimensional solution- and solid-state structures of the human immunodeficiency virus type-1 (HIV-1) matrix protein have been determined recently in our laboratories by NMR and X-ray crystallographic methods (Massiah et al. 1994. J Mol Biol 244:198-223; Hill et al. 1996. Proc Natl Acad Sci USA 93:3099-3104). The matrix protein exists as a monomer in solution at low millimolar protein concentrations, but forms trimers in three different crystal lattices. Although the NMR and X-ray structures are similar, detailed comparisons have revealed an approximately 6 A displacement of a short 3(10) helix (Pro 66-Gly 71) located at the trimer interface. High quality electron density and nuclear Overhauser effect (NOE) data support the integrity of the X-ray and NMR models, respectively. Because matrix apparently associates with the viral membrane as a trimer, displacement of the 3(10) helix may reflect a physiologically relevant conformational change that occurs during virion assembly and disassembly. These findings further suggest that Pro 66 and Gly 71, which bracket the 3(10) helix, serve as "hinges" that allow the 3(10) helix to undergo this structural reorientation.
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PMID:Comparison of the NMR and X-ray structures of the HIV-1 matrix protein: evidence for conformational changes during viral assembly. 897 48

The Nef protein alters T cell receptor (TCR) signaling in T cells and is critical for the pathogenesis of AIDS. We used a transient expression assay in a human CD4+ T cell line to analyze the interaction of Nef with the TCR machinery. We show that, in addition to down-regulating CD4 expression on the cell surface, Nef blocks a receptor-proximal event in CD3 signaling. Analysis of a large number of mutant Nef proteins demonstrated that the effects of Nef on CD4 expression and on CD3 signaling are separable. The ability of Nef to block CD3 signaling was selectively abolished by mutations in the central part of the Nef protein and in particular by those known to disrupt the SH3 binding surface in the structured core of Nef. In contrast, the ability of Nef to down-regulate CD4 expression was selectively abolished by two clusters of mutations, one in the N-terminal and one in the C-terminal region of Nef. These two regions correspond to the two flexible loops in Nef as predicted by solution NMR analysis. We show that this general functional organization is conserved between the Nef proteins of the human and simian immunodeficiency viruses (HIV-1 and SIV). Our data demonstrate that Nef has at least two independent mechanisms to alter TCR function and thus may interfere with a range of T cell responses.
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PMID:Separable functions of Nef disrupt two aspects of T cell receptor machinery: CD4 expression and CD3 signaling. 904 97


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