Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.5.1.18 (glutathione S-transferase)
22,582 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Tat protein, a transactivating factor of the human immunodeficiency virus type I, acts also as an extracellular molecule. Heparin affects the bioavailability and biological activity of extracellular Tat (Rusnati, M., Coltrini, D., Oreste, P., Zoppetti, G., Albini, A., Noonan, D., D'Adda di Fagagna, F., Giacca, M., and Presta, M. (1997) J. Biol. Chem. 272, 11313-11320). Here, a series of homogeneously sized, (3)H-labeled heparin fragments were evaluated for their capacity to bind to free glutathione S-transferase (GST)-Tat protein and to immobilized GST-Tat. Hexasaccharides represent the minimum sized heparin fragments able to interact with GST-Tat at physiological ionic strength. Also, the affinity of binding increases with increasing the molecular size of the oligosaccharides, with large fragments (>/=18 saccharides) approaching the affinity of full-size heparin. 6-Mer heparin binds GST-Tat with a dissociation constant (K(d)) equal to 0.7 +/- 0.4 microM and a molar oligosaccharide:GST-Tat ratio of about 1:1. Interaction of GST-Tat with 22-mer or full-size heparin is consistent instead with two-component binding. At subsaturating concentrations, a single molecule of heparin interacts with 4-6 molecules of GST-Tat with high affinity (K(d) values in the nanomolar range of concentration); at saturating concentrations, heparin binds GST-Tat with lower affinity (K(d) values in the micromolar range of concentration) and a molar oligosaccharide:GST-Tat ratio of about 1:1. In agreement with the binding data, a positive correlation exists between the size of heparin oligosaccharides and their capacity to inhibit cell internalization, long terminal repeat-transactivating activity of extracellular Tat in HL3T1 cells, and its mitogenic activity in murine adenocarcinoma T53 Tat-less cells. The data demonstrate that the modality of heparin-Tat interaction is strongly affected by the size of the saccharide chain. The possibility of establishing multiple interactions increases the affinity of large heparin fragments for Tat protein and the capacity of the glycosaminoglycan to modulate the biological activity of extracellular Tat.
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PMID:Multiple interactions of HIV-I Tat protein with size-defined heparin oligosaccharides. 1049 73

Bruton's tyrosine kinase (Btk) is considered an essential signal transducer in B-cells. Mutational defects are associated with a severe immunodeficiency syndrome, X-chromosome linked agammaglobulinemia (XLA). Here we show by coimmunoprecipitation that a member of the protein kinase C (PKC) family, PKCmu, is constitutively associated with Btk. Neither antigen receptor (Ig) crosslinking nor stimulation of B-cells with phorbol ester or H(2)O(2) affected Btk/PKCmu interaction. GST precipitation analysis revealed association of the Btk pleckstrin/Tec homology domain with PKCmu. Transient overexpression of PKCmu deletion mutants as well as expression of selected PKCmu domains in 293T cells revealed that both the kinase domain and the regulatory C1 region are independently capable of binding to the Btk PH-TH domain. These data show the existence of a PKCmu/Btk complex in vivo and identify two PKCmu domains that participate in Btk interaction.
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PMID:Bruton's tyrosine kinase (Btk) associates with protein kinase C mu. 1056 98

We have recently demonstrated that simian immunodeficiency virus (SIV) Nef binds to the zeta chain of the T-cell receptor (TCR), leading to its down-modulation from T-cell surfaces (I. Bell, C. Ashman, J. Maughan, E. Hooker, F. Cook, and T. A. Reinhart, J. Gen. Virol. 79:2717-2727, 1998). Using a panel of human as well as rhesus macaque TCR zeta cytoplasmic domain mutants, we have identified in this report two linear peptides in the cytoplasmic domain of TCR zeta which independently interact with SIV Nef. Each SIV Nef interaction domain was sufficient in the absence of the other for interaction with SIV Nef in a yeast two-hybrid assay. In parallel, we demonstrated that Nef down-modulation of CD8-TCR zeta fusion proteins containing full-length or truncated portions of the TCR zeta cytoplasmic domain occurs in transiently transfected 293T cells. Furthermore, using proteins expressed in Escherichia coli, a glutathione S-transferase-Nef fusion protein coprecipitated histidine-tagged portions of the TCR zeta cytoplasmic domain which contained SNID-1 or SNID-2. The peptides targeted by SIV Nef, YNELNL and YSEIGMKGERRR, are portions of the first and second of three immunoreceptor tyrosine-based activation motifs which are important in signal transduction, thymocyte development, and TCR biogenesis. These results demonstrate that SIV Nef binds to two distinct domains on TCR zeta in the absence of other T-cell-specific factors, and that interaction with either domain is sufficient to cause down-modulation of TCR zeta.
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PMID:The T-cell receptor zeta chain contains two homologous domains with which simian immunodeficiency virus Nef interacts and mediates down-modulation. 1070 44

We previously demonstrated that an envelope mutant of human immunodeficiency virus type 1 lacking the entire cytoplasmic domain interferes in trans with the production of infectious virus by inclusion of the mutant envelope into the wild-type envelope complex. We also showed that the envelope incorporation into virions is not affected when the wild-type envelope is coexpressed with the mutant envelope. These results suggest that an oligomeric structure of the cytoplasmic domain is functionally required for viral infectivity. To understand whether the cytoplasmic domain of human immunodeficiency virus type 1 transmembrane protein gp41 has the potential to self-assemble as an oligomer, in the present study we fused the coding sequence of the entire cytoplasmic domain at 3' to the Escherichia coli malE gene, which encodes a monomeric maltose-binding protein. The expressed fusion protein was examined by chemical cross-linking, sucrose gradient centrifugation, and gel filtration. The results showed that the cytoplasmic domain of gp41 assembles into a high-ordered structural complex. The intersubunit interaction of the cytoplasmic domain was also confirmed by a mammalian two-hybrid system that detects protein-protein interactions in eucaryotic cells. A cytoplasmic domain fragment expressed in eucaryotic cells was pulled down by glutathione-Sepharose 4B beads via its association with another cytoplasmic domain fragment fused to the C terminus of the glutathione S-transferase moiety. We also found that sequences encompassing the lentiviral lytic peptide-1 and lentiviral lytic peptide-2, which are located within residues 828-856 and 770-795, respectively, play a critical role in cytoplasmic domain self-assembly. Taken together, the results from the present study indicate that the cytoplasmic domain of gp41 by itself is sufficient to assemble into a multimeric structure. This finding supports the hypothesis that a multimeric form of the gp41 cytoplasmic domain plays a crucial role in virus infectivity.
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PMID:Multimerization potential of the cytoplasmic domain of the human immunodeficiency virus type 1 transmembrane glycoprotein gp41. 1074 37

The human immunodeficiency virus type 1 (HIV-1) RNA-binding Rev protein governs the expression of structural and enzymatic viral proteins at a posttranscriptional level. Binding of Rev to the stem-loop IIB (SLIIB) sequence of the Rev-response element (RRE) within unspliced and singly spliced viral mRNAs and to the nuclear export signal-binding receptor, hCRM1 (or exportin 1), is required for the export of these transcripts to the cytoplasm. We have previously shown that herpes simplex virus type 1 (HSV-1) RNA-binding Us11 protein is able to bind the RRE and substitute for Rev in inducing the expression of HIV-1 envelope glycoproteins. We show here that Us11 cannot substitute for Rev in rescuing a rev-deleted HIV-1 provirus. However, HIV-1 production is observed when Us11 is expressed with suboptimal amounts of Rev. An in vivo RNA-protein binding assay indicates that Us11 is unable to directly interact with the SLIIB RNA but can bind Rev assembled on that stem-loop structure. This association of US11 with Rev, which was confirmed by in vivo coimmunoprecipitation and GST-pulldown assays, therefore underlies a biological Us11-Rev cooperation. Furthermore this cooperation was shown to remain susceptible to the effect of leptomycin B, which blocks the binding of hCRM1 to the nuclear export signal of Rev. These observations performed with intron-containing constructs provide evidence that HSV-1 Us11 protein is not directly involved in the cytoplasmic accumulation of viral mRNAs but may be rather acting as an auxiliary protein, thus allowing this retroviral protein to fulfill the nuclear export of these transcripts and to rescue HIV-1 production.
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PMID:The herpes simplex virus 1 Us11 protein cooperates with suboptimal amounts of human immunodeficiency virus type 1 (HIV-1) Rev protein to rescue HIV-1 production. 1077 78

Vpx is a virion-associated protein of human immunodeficiency virus type 2 (HIV-2) and simian immunodeficiency viruses. The yeast two-hybrid system was used to identify invariant chain (Ii) as a cellular protein that interacts with HIV-2 Vpx. Vpx-Ii interaction was confirmed in cell-free reactions using bacterially expressed glutathione S-transferase fusion proteins and by coimmunoprecipitation in transfected and infected cells. In chronically infected cells expressing Vpx, Ii levels were markedly decreased, presumably due to enhanced degradation. These findings suggest that Vpx may disrupt major histocompatibility complex class II antigen presentation.
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PMID:Interaction of human immunodeficiency virus type 2 Vpx and invariant chain. 1084 1

The virus infectivity factor (Vif) protein facilitates the replication of human immunodeficiency virus type 1 (HIV-1) in primary lymphocytes and macrophages. Its action is strongly dependent on the cellular environment, and it has been proposed that the Vif protein counteracts cellular activities that would otherwise limit HIV-1 replication. Using a glutathione S-transferase pull-down assay, we identified that Vif binds specifically to the Src homology 3 domain of Hck, a tyrosine kinase from the Src family. The interaction between Vif and the full-length Hck was further assessed by co-precipitation assays in vitro and in human cells. The Vif protein repressed the kinase activity of Hck and was not itself a substrate for Hck phosphorylation. Within one single replication cycle of HIV-1, Hck was able to inhibit the production and the infectivity of vif-deleted virus but not that of wild-type virus. Accordingly, HIV-1 vif- replication was delayed in Jurkat T cell clones stably expressing Hck. Our data demonstrate that Hck controls negatively HIV-1 replication and that this inhibition is suppressed by the expression of Vif. Hck, which is present in monocyte-macrophage cells, represents the first identified cellular inhibitor of HIV-1 replication overcome by Vif.
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PMID:The tyrosine kinase Hck is an inhibitor of HIV-1 replication counteracted by the viral vif protein. 1127 65

Extracellular Tat protein, the transactivating factor of the human immunodeficiency virus type 1 (HIV-1), modulates gene expression, growth, and angiogenic activity in endothelial cells by interacting with the vascular endothelial growth factor (VEGF) receptor-2 (Flk-1/KDR). Recombinant Tat protein, produced as glutathione-S-transferase chimera (GST-Tat), activates mitogen-activated protein kinase (MAPK) ERK(1/2) in human, murine, and bovine endothelial cells whereas GST is ineffective. In bovine aortic endothelial cells, GST-Tat and the 165 amino acid VEGF isoform (VEGF165) induce transient ERK(1/2) phosphorylation with similar potency and kinetics. The synthetic peptide Tat(41-60), but not peptides Tat(1-21) and Tat(71-86), causes ERK(1/2) phosphorylation, thus implicating Tat/KDR interaction in the activation of this signalling pathway. Accordingly, GST-Tat induces ERK(1/2) phosphorylation in KDR-transfected porcine aortic endothelial cells but not in parental cells. MAPK kinase inhibitors PD098059 and U0126 prevent ERK(1/2) phosphorylation by Tat. However, they do not affect the angiogenic activity exerted by Tat in the murine Matrigel plug and chick embryo chorioallantoic membrane assays. Blocking of MAPK kinase activity impairs instead the angiogenic response to VEGF165 and to fibroblast growth factor-2 (FGF-2). Our data demonstrate that ERK(1/2) activation following the interaction of HIV-1 Tat protein with endothelial cell Flk-1/KDR receptor does not represent an absolute requirement for a full angiogenic response to this growth factor that appears to utilize mechanism(s) at least in part distinct from those triggered by other prototypic angiogenic growth factors.
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PMID:Activation of endothelial cell mitogen activated protein kinase ERK(1/2) by extracellular HIV-1 Tat protein. 1140 52

Pur alpha is a highly conserved, eukaryotic sequence-specific DNA- and RNA-binding protein involved in diverse cellular and viral functions including transcription, replication, and cell growth. Pur alpha exerts its activity in part by interacting with other viral and cellular proteins. One such protein is the human immunodeficiency virus (HIV) type I regulatory protein Tat. Earlier studies have demonstrated that this interaction is mediated by Pur alpha-associated RNA (PARNA) and that RNA immunopurified from mammalian expressed Pur alpha was capable of reconstituting the interaction between these two proteins. In the current study, we characterize four RNA species which were immunopurified with Pur alpha. Northern blot analysis with one of the PARNAs revealed a highly abundant signal of approximately 2.0 kilobases (kb) present in all cell lines tested. Sequence analysis of each of the four PARNA clones revealed a high homology to different regions of the human 18S ribosomal RNA sequence. Based on this homology, we investigated the influence of Pur alpha on translation. Luciferase assays were performed after coupled in vitro transcription/translation reactions with a vector containing a luciferase reporter construct and increasing concentrations of BSA, GST, and GST-Pur alpha. Inclusion of GST-Pur alpha in these reactions resulted in a dose-dependent inhibition of luciferase activity. Similar inhibition was observed with in vitro translation reactions performed with in vitro transcribed luciferase RNA and increasing concentrations of GST-Pur alpha. In control experiments, inclusion of increasing concentrations of GST-Pur alpha with luciferase protein resulted in no effect on luciferase activity. Taken together, these data demonstrate that Pur alpha inhibits translation reactions in vitro. Moreover, this Pur alpha-mediated inhibition of translation can be abrogated by HIV-1 Tat protein.
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PMID:Single-stranded nucleic acid-binding protein, Pur alpha, interacts with RNA homologous to 18S ribosomal RNA and inhibits translation in vitro. 1159 4

Common variable immunodeficiency (CVI) patients are at high relative risk of developing non-Hodgkin lymphomas (NHL), mainly represented by B-lineage diffuse large cell lymphomas. The molecular pathogenesis and histogenesis of CVI-related NHL are poorly understood. We have thus attempted to provide a detailed molecular characterization of their histogenesis and pathogenesis. A panel of 5 CVI-related NHL was subjected to detailed analysis of histogenetic markers (mutations of immunoglobulin variable heavy chain-IgVH and of BCL-6 genes) acquired by B-cells at the time of germinal center transit. Somatic hypermutation of IgVH and BCL-6 genes occurred in 5/5 cases; in all cases, mutations were stable with no evidence of ongoing mutation processes. In 3/5 cases, the pattern of IgVH mutations was consistent with selection and stimulation of the tumor clone by antigen. To further clarify the pathogenesis, samples were tested for inactivation by promoter hypermethylation of the genes 0(6)-methylguanine-DNA-methyltransferase (MGMT) and glutathione S-transferase (GST) p1, which code for detoxifying enzymes, as well as of death-associated protein (DAP)-kinase, coding for a proapoptotic molecule. Promoter hypermethylation of MGMT, GSTp1 and DAP-kinase was detected in 2/5, 3/5 and 3/5 CVI-related NHL, respectively. Overall, these data indicate that: i) similarly to other immunodeficiency-related NHL, CVI-related NHL derive from germinal center-related B-cells, namely centrocytes or post-germinal center B-cells; ii) antigen stimulation and selection are involved in the development of at least a fraction of these cases; iii) hypermethylation of the MGMT, DAP-kinase and GSTp1 genes occurs at sustained frequencies in CVI-related NHL and may provide novel prognostic markers and therapeutic targets for the clinical management of these lymphomas.
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PMID:Molecular characterization of common variable immunodeficiency-related lymphomas. 1169 5


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