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)

Proteomics is a relatively new approach for understanding the pathology and pathogenesis of various diseases. It has also been used for characterizing the modifications in protein expression during the development of interstitial lung diseases, in lung tumors, or following exposure to exogenous stress signals. We compared the protein composition of primary human lung fibroblasts derived from patients with lung fibrosis and control fibroblasts of unaffected lung tissues. We found a predominant modulation in proteins related to the cytoskeleton, including decreased expression of vimentin and lamin A/C, and increased expression of moesin. Furthermore, we observed lower levels of components of the antioxidative system, such as omega class glutathione S-transferase and an up-regulation of an intracellular chloride channel. In fibroblasts obtained from fibrotic lungs, the expression of a major histocompatibility complex class I C was decreased, and so was the expression of tripeptidyl-peptidase-I-precursor, a collagen-degrading exopeptidase. Our results and the studies reviewed in this paper represent just the beginning of detailed studies that should unravel the relevance and the functional consequences of differential protein expressions in the diseased lung.
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PMID:Clinical proteomics in lung diseases. 1523 23

Renal tubular epithelial cells in all nephron segments express a distinct member of the metalloprotease-disintegrin family, ADAM9 (a disintegrin and metalloprotease 9), in a punctate basolateral distribution co-localized to the beta1 integrin chain [Mahimkar, Baricos, Visaya, Pollock and Lovett (2000) J. Am. Soc. Nephrol. 11, 595-603]. Discrete segments of the nephron express several defined beta1 integrins, suggesting that ADAM9 interacts with multiple renal integrins and thereby regulates epithelial cell-matrix interactions. Intact ADAM9 and a series of deletion constructs sequentially lacking the metalloprotease domain and the disintegrin domain were assembled as chimaeras with a C-terminal GFP (green fluorescent protein) tag. Stable expression of the ADAM9/GFP protein on the surface of HEK-293 cells (human embryonic kidney 293 cells) significantly decreased adhesion to types I and IV collagen, vitronectin and laminin, but had little effect on adhesion to fibronectin. Expression of the disintegrin/cysteine-rich/GFP construct yielded a similar, but more marked pattern of decreased adhesion. Expression of the cysteine-rich/GFP construct had no effect on adhesion, indicating that the disintegrin domain was responsible for the competitive inhibition of cell-matrix binding. To define the specific renal tubular beta1 integrins interacting with the ADAM9 disintegrin domain, a recombinant GST (glutathione S-transferase)-disintegrin protein was used as a substrate in adhesion assays in the presence or absence of specific integrin-blocking antibodies. Inclusion of antibodies to alpha1, alpha3, alpha6, alphav and beta1 blocked adhesion of HEK-293 cells to GST-disintegrin protein. Immobilized GST-disintegrin domain perfused with renal cortical lysates specifically recovered the alpha3, alpha6, alphav and beta1 integrin chains by Western analysis. It is concluded that ADAM9 is a polyvalent ligand, through its disintegrin domain, for multiple renal integrins of the beta1 class.
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PMID:The disintegrin domain of ADAM9: a ligand for multiple beta1 renal integrins. 1536 Oct 64

Integrin receptors are crucial players in cell adhesion and migration. Identification and characterization of cellular proteins that interact with their short alpha and beta cytoplasmic tails will help to elucidate the molecular mechanisms by which integrins mediate bi-directional signaling across the plasma membrane. Integrin alpha2beta1 is a major collagen receptor but to date, only few proteins have been shown to interact with the alpha2 cytoplasmic tail or with the alpha2beta1 complex. In order to identify novel binding partners of a alpha2beta1cytoplasmic domain complex, we have generated recombinant GST-fusion proteins, incorporating the leucine zipper heterodimerization cassettes of Jun and Fos. To ascertain proper functionality of the recombinant proteins, interaction with natural binding partners was tested. GST-alpha2 and GST-Jun alpha2 bound His-tagged calreticulin while GST-beta1 and GST-Fos beta1 proteins bound talin. In screening assays for novel binding partners, the immobilized GST-Jun alpha2/GST-Fos beta1 heterodimeric complex, but not the single subunits, interacted specifically with endothelial cell-derived vimentin. Vimentin, an abundant intermediate filament protein, has previously been shown to co-localize with alphavbeta3-positive focal contacts. Here, we provide evidence that this interaction also occurs with alpha2beta1-enriched focal adhesions and we further show that this association is lost after prolonged adhesion of endothelial cells to collagen.
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PMID:The intermediate filament protein vimentin binds specifically to a recombinant integrin alpha2/beta1 cytoplasmic tail complex and co-localizes with native alpha2/beta1 in endothelial cell focal adhesions. 1577 92

Collagen type II is an extracellular matrix protein important for cartilage and bone formation, and its expression is controlled by multiple cis- and trans-acting elements, including the zinc finger transcription factor alpha A-crystallin-binding protein 1 (CRYBP1). Here we show that MSX2-interacting nuclear target protein (MINT), a conserved transcriptional repressor, associates with CRYBP1 and negatively regulates the transactivation of the collagen type II gene (Col2a1) enhancer. We identified CRYBP1 as a binding partner of MINT by screening a mouse embryonic cDNA library using the yeast two-hybrid system. We demonstrated that the C terminus of MINT interacts with the C terminus of CRYBP1 using the mammalian cell two-hybrid assay, glutathione S-transferase pull-down, and co-immunoprecipitation analyses. Furthermore, MINT and CRYBP1 form a complex on the Col2a1 enhancer, as shown by chromatin immunoprecipitation and gel shift assays. In the presence of CRYBP1, overexpression of MINT or its C-terminal fragment in cells repressed a reporter construct driven by the Col2a1 enhancer elements. This transcription repression is dependent on histone deacetylase, the main co-repressor recruited by MINT. The present study shows that MINT is involved in CRYBP1-mediated Col2a1 gene repression and may play a role in regulation of cartilage development.
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PMID:Mint represses transactivation of the type II collagen gene enhancer through interaction with alpha A-crystallin-binding protein 1. 1577 99

CD93 is a cell-surface glycoprotein that has been shown to influence defence collagen-enhanced Fc-receptor or CR1-mediated phagocytosis of suboptimally opsonized targets in vitro, and CD93-deficient mice are defective in the clearance of apoptotic cells in vivo. To investigate the mechanism of CD93 modulation of phagocytic activity, GST fusion proteins containing the 47 amino acid intracellular domain (GST-Cyto), or various mutants of the intracellular domain of CD93, were constructed and used to identify intracellular CD93-binding molecules. The intracellular protein moesin, well characterized for its role in linking transmembrane proteins to the cytoskeleton and in cytoskeletal remodelling, bound to GST-Cyto when either cell lysates or recombinant moesin were used as a source of interacting molecules. An association of moesin with CD93 within intact cells was confirmed by co-capping moesin with CD93 in human monocytes. The moesin-binding site on CD93 mapped to the first four positively charged amino acids in the juxtamembrane region of the CD93 cytoplasmic tail. Interestingly, deletion of the last 11 amino acids from the C terminus of CD93 (GST-Cyto-C11) dramatically increased moesin binding to the cytoplasmic tail of CD93 in the cell lysate assay, but not when the binding of purified recombinant moesin was assessed. Furthermore, moesin binding to CD93 was enhanced by the addition of phosphatidylinositol 4,5-bisphosphate (PIP(2)). Taken together, these data suggest that the interaction of moesin with the CD93 cytoplasmic domain is modulated by binding of other intracellular molecules to the C11 region and implies that a PIP(2) signalling pathway is involved in CD93 function.
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PMID:Modulated interaction of the ERM protein, moesin, with CD93. 1581 98

The eukaryotic initiation factor 5A (eIF5A), a factor essential for eukaryotic cell proliferation, is the only cellular protein containing the polyamine-derived amino acid hypusine [N(epsilon)-(4-amino-2-hydroxybutyl)lysine]. Hypusine is formed in a posttranslational modification that involves two sequential enzymatic steps catalyzed by deoxyhypusine synthase and deoxyhypusine hydroxylase (DOHH). By screening a Saccharomyces cerevisiae GST-ORF library for expression of DOHH activity, we have cloned YJR070C as the gene encoding DOHH and identified the human homolog DOHH gene, HLRC1. Purified recombinant yeast and human DOHH enzymes effectively catalyzed hydroxylation of the deoxyhypusine residue in the eIF5A intermediate. Overexpression of human DOHH along with eIF5A precursor and deoxyhypusine synthase was required for overproduction of mature, hypusine-containing eIF5A in 293T and other mammalian cells. The Saccharomyces cerevisiae strain with deletion of YJR070C contained only deoxyhypusine but no hypusine, indicating that YJR070C was the single DOHH gene in this organism. One highly conserved DOHH homolog gene is found in a variety of eukaryotes from yeast to human. Sequence and structural analyses reveal that DOHH belongs to a family of HEAT-repeat-containing proteins, consisting of eight tandem repeats of an alpha-helical pair (HEAT motif) organized in a symmetrical dyad. The predicted structure is unrelated to the double-stranded beta-helix type structures of the Fe(II)- and 2-oxoacid-dependent dioxygenases, such as collagen prolyl or lysyl hydroxylases. However, metal coordination sites composed of four strictly conserved histidine-glutamate sequences were identified, suggesting that DOHH enzymes have convergently evolved an iron-dependent hydroxylation mechanism.
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PMID:Molecular cloning, expression, and structural prediction of deoxyhypusine hydroxylase: a HEAT-repeat-containing metalloenzyme. 1637 67

Visualization of the formation and orientation of collagen fibers in tissue engineering experiments is crucial for understanding the factors that determine the mechanical properties of tissues. In this study, collagen-specific fluorescent probes were developed using a new approach that takes advantage of the inherent specificity of collagen binding protein domains present in bacterial adhesion proteins (CNA35) and integrins (GST-alpha1I). Both collagen binding domains were obtained as fusion proteins from an Escherichia coli expression system and fluorescently labeled using either amine-reactive succinimide (CNA35) or cysteine-reactive maleimide (GST-alpha1I) dyes. Solid-phase binding assays showed that both protein-based probes are much more specific than dichlorotriazinyl aminofluorescein (DTAF), a fluorescent dye that is currently used to track collagen formation in tissue engineering experiments. The CNA35 probe showed a higher affinity for human collagen type I than did the GST-alpha1I probe (apparent K(d) values of 0.5 and 50 microM, respectively) and showed very little cross-reactivity with noncollagenous extracellular matrix proteins. The CNA35 probe was also superior to both GST-alpha1I and DTAF in visualizing the formation of collagen fibers around live human venous saphena cells. Immunohistological experiments on rat tissue showed colocalization of the CNA35 probe with collagen type I and type III antibodies. The fluorescent probes described here have important advantages over existing methods for visualization of collagen, in particular for monitoring the formation of collagen in live tissue cultures over prolonged time periods.
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PMID:Fluorescently labeled collagen binding proteins allow specific visualization of collagen in tissues and live cell culture. 1647 6

A collagen-degrading thermophile, Geobacillus collagenovorans MO-1, extracellularly produces a collagenolytic protease with a large molecular mass. Complete nucleotide sequencing of this gene after gene cloning revealed that the collagenolytic protease is a member of the subtilisin family of serine proteases and consists of a signal sequence for secretion, a prosequence for maturation, a catalytic region, 14 direct repeats of 20 amino acids at the C terminus, and a region with unknown function intervening between the catalytic region and the numerous repeats. Since the unusual repeats are most likely to be cleaved in the secreted form of the enzyme, the intervening region was investigated to determine whether it participates in collagen binding to facilitate collagen degradation. It was found that the mature collagenolytic protease containing the intervening region at the C terminus bound collagen but not the other insoluble proteins, elastin and keratin. Furthermore, the intervening region fused with glutathione S-transferase showed a collagen-binding ability comparable to that of the mature collagenolytic protease. The collagen-binding ability was finally attributed to two-thirds of the intervening region which is rich in beta-strands and is approximately 35 kDa in molecular mass. In the collagenolytic protease from strain MO-1, hydrogen bonds most likely predominate over the hydrophobic interaction for collagen binding, since a higher concentration of NaCl released collagen from the enzyme surface but a nonionic detergent could not. To the best of our knowledge, this is the first report of a thermophilic collagenolytic protease containing the collagen-binding segment.
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PMID:Characteristic features in the structure and collagen-binding ability of a thermophilic collagenolytic protease from the thermophile Geobacillus collagenovorans MO-1. 1695 49

Nuclear receptors such as constitutive androstane receptor (CAR) and pregnane X receptor (PXR) regulate the transcription of cytochromes P450 and transporters. We investigated whether quantitative and functional changes in CAR and PXR could affect bilirubin detoxification in chronic arthritis. The CAR mRNA level was significantly decreased in the liver of mice with collagen-induced arthritis (CIA) compared with control mice. In normal mice treated with CAR agonists, relatively rapid elimination of bilirubin was observed after its intravenous injection. Next, we investigated the effects of CAR on bilirubin-detoxifying enzymes and transporters in arthritis. The mRNA levels of organic anion transporter peptide (OATP) 2, glutathione S-transferase (GST) A1, and GSTA2 were decreased in CIA mice, whereas the mRNA levels of OATP4, UDP-glucuronosyl-transferase 1A1, and multidrug resistance-associated protein 2 remained unchanged. The protein levels and transport activities of OATP2 were also decreased in CIA mice. Furthermore, the CIA mice actually exhibited retarded elimination of bilirubin after its intravenous injection. These results indicate that alterations to CAR during arthritis affect the elimination of bilirubin because of changes in multiple bilirubin-detoxifying enzymes and transporters.
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PMID:Effects of alterations in CAR on bilirubin detoxification in mouse collagen-induced arthritis. 1710 61

The protein bullous pemphigoid antigen-2 (BPAG2/BP180/collagen type XVII) plays a key role in attachment of basal keratinocytes to epidermal basement membrane. The binding of BP180 with either integrin alpha6, integrin beta4, or bullous pemphigoid antigen-1 (BPAG1/BP230) is critical for this attachment in skin. The protein 14-3-3 sigma, also known as stratifin and a marker for epithelial cells, is a member of a highly conserved small acidic 14-3-3 protein family naturally found in all eukaryotic cells. Here, we have used a 14-3-3sigma GST pull-down screening assay and showed that sigma (sigma) isoform of the 14-3-3 protein family interacts with the cytoplasmic N-terminal domain of BP180. Analysis of a series of truncated or deleted 14-3-3sigma revealed that only intact 14-3-3sigma molecule, but not any of its fragments can interact with BP180. This finding suggests that conformation and possible dimerization of 14-3-3 sigma is essential for this interaction. Further, a BP180 co-immunoprecipitation (IP) and its reverse IP assays were conducted and the results confirmed that 14-3-3 sigma interacts with cytoplasmic domain, but not ecto-domain of the BP180. In conclusion, the finding of this study provides evidence that 14-3-3sigma isoform interacts with BP180 which is a major component of hemidesmosome involved in the attachment of epidermis to the basement membrane in skin. However, the significance of this interaction in hemidesmosome formation and/or attachment needs to be explored.
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PMID:14-3-3 sigma isoform interacts with the cytoplasmic domain of the transmembrane BP180 in keratinocytes. 1744 72


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