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)

Solvent-induced equilibrium unfolding of a homodimeric class sigma glutathione transferase (GSTS1-1, EC 2.5.1.18) was characterized by tryptophan fluorescence, anisotropy, enzyme activity, 8-anilino-1-naphthalenesulfonate (ANS) binding, and circular dichroism. Urea induces a triphasic unfolding transition with evidence for two well-populated thermodynamically stable intermediate states of GSTS1-1. The first unfolding transition is protein concentration independent and involves a change in the subunit tertiary structure yielding a partially active dimeric intermediate (i.e., N2 left and right arrow I2). This is followed by a protein concentration dependent step in which I2 dissociates into compact inactive monomers (M) displaying enhanced hydrophobicity. The third unfolding transition, which is protein concentration independent, involves the complete unfolding of the monomeric state. Increasing NaCl concentrations destabilize N2 and appear to shift the equilibrium toward I2 whereas the stability of the monomeric intermediate M is enhanced. The binding of substrate or product analogue (i.e., glutathione or S-hexylglutathione) to the protein's active site stabilizes the native dimeric state (N2), causing the first two unfolding transitions to shift toward higher urea concentrations. The stability of M was not affected. The data implicate a region at/near the active site in domain I (most likely alpha-helix 2) as being highly unstable/flexible which undergoes local unfolding, resulting initially in I2 formation followed by a disruption in quaternary structure to a monomeric intermediate. The unfolding/refolding pathway is compared with those observed for other cytosolic GSTs and discussed in light of the different structural features at the subunit interfaces, as well as the evolutionary selection of this GST as a lens crystallin.
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PMID:Class sigma glutathione transferase unfolds via a dimeric and a monomeric intermediate: impact of subunit interface on conformational stability in the superfamily. 979 17

The soluble S-crystallin constitutes the major lens protein in cephalopods. The primary amino acid sequence of S-crystallin shows an overall 41% identity with the digestive gland sigma-class glutathione transferase (GST) of cephalopod. However, the lens S-crystallin fails to bind to the S-hexylglutathione affinity column and shows very little GST activity in the nucleophilic aromatic substitution reaction between GSH and 1-chloro-2,4-dinitrobenzene. When compared with other classes of GST, the S-crystallin has an 11-amino acid residues insertion between the conserved alpha4 and alpha5 helices. Based on the crystal structure of squid sigma-class GST, a tertiary structure model for the octopus lens S-crystallin is constructed. The modeled S-crystallin structure has an overall topology similar to the squid sigma-class GST, albeit with longer alpha4 and alpha5 helical chains, corresponding to the long insertion. This insertion, however, makes the active center region of S-crystallin to be in a more closed conformation than the sigma-class GST. The active center region of S-crystallin is even more shielded and buried after dimerization, which may explain for the failure of S-crystallin to bind to the immobilized-glutathione in affinity chromatography. In the active site region, the electrostatic potential surface calculated from the modeled structure is quite different from that of squid GST. The positively charged environment, which contributes to stabilize the negatively charged Meisenheimer complex, is altered in S-crystallin probably because of mutation of Asn99 in GST to Asp101 in S-crystallin. Furthermore, the important Phe106 in authentic GST is changed to His108 in S-crystallin. Combining the topological differences as revealed by computer graphics and sequence variation at these structurally relevant residues provide strong structural evidences to account for the much decreased GST activity of S-crystallin as compared with the authentic GST of the digestive gland.
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PMID:Homology modeling of cephalopod lens S-crystallin: a natural mutant of sigma-class glutathione transferase with diminished endogenous activity. 992 73

Protein S-thiolation is a process in which under oxidative stress, vulnerable sulfhydryl groups of proteins are conjugated to non-protein thiols such as glutathione (GSH) or cysteine resulting in the formation of protein-thiol mixed disulfides, protein-S-S-glutathione (PSSG) and protein-S-S-cysteine (PSSC). This process spontaneously disrupts the redox homeostasis of the cells, which in turn leads to functional disturbances in the respective tissue. In the ocular lens, such modification of proteins may trigger a cascade of events starting with the alteration of protein conformation, protein/enzyme deactivation, protein-S-S-protein aggregation and eventually lens opacification or cataract. Generally, the first line of defense system in the cells protects the lens proteins against such damage. Recent studies in our laboratory have shown that in addition to this defense system, lens cells also possess a well developed system to repair the oxidative damage to the lens proteins. We have identified this repair system as thioltransferase (TTase) and have proved that TTase by its dethiolase activity reverses the protein S-thiolation process which returns the oxidatively damaged lens proteins/enzymes to their original reduced state and restores their physiological functions. We investigated if this repair mechanism was mediated by enzymes other than TTase. We studied glutathione S-transferase (GST) and report here for the first time the cloning, high level expression, and purification of human lens mu and pi isoforms of GST. A comparative study of recombinant human lens TTase and GST (mu and pi) on their dethiolating abilities using lens crystallin-thiol mixed disulfides showed that the lens TTase is 60-70% more efficient in the dethiolation/repair process than GST. When TTase and GST were tested in conjunction for the dethiolation of thiol mixed disulfides, there was no significant enhancement of dethiolase activity. These findings suggest that TTase by itself is an efficient enzyme in the dethiolation/repair process and hence can be considered a crucial system to counteract oxidative stress in the lens.
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PMID:Does glutathione-S-transferase dethiolate lens protein-thiol mixed disulfides?-A comparative study with thioltransferase. 1037 35

S-Crystallin from octopus lens has a tertiary structure similar to sigma-class glutathione transferase (GST). However, after isolation from the lenses, S-crystallin was found to aggregate more easily than sigma-GST. In vitro experiments showed that the lens S-crystallin can be polymerized and finally denatured at increasing concentration of urea or guanidinium chloride (GdmCl). In the intermediate concentrations of urea or GdmCl, the polymerized form of S-crystallin is aggregated, as manifested by the increase in light scattering and precipitation of the protein. There is a delay time for the initiation of polymerization. Both the delay time and rate of polymerization depend on the protein concentration. The native protein showed a maximum fluorescence emission spectrum at 341 nm. The GdmCl-denatured protein exhibited two fluorescence maxima at 310 nm and 358 nm, respectively, whereas the urea-denatured protein showed a fluorescence peak at 358 nm with a small peak at 310 nm. The fluorescence intensity was quenched. Monomers, dimers, trimers, and polymers of the native protein were observed by negative-stain electron microscopic analysis. The aggregated form, however, showed irregular structure. The aggregate was solubilized in high concentrations of urea or GdmCl. The redissolved denatured protein showed an identical fluorescence spectrum to the protein solution that was directly denatured with high concentrations of urea or GdmCl. The denatured protein was readily refolded to its native state by diluting with buffer solution. The fluorescence spectrum of the renatured protein solution was similar to that of the native form. The phase diagrams for the S-crystallin in urea and GdmCl were constructed. Both salt concentration and pH value of the solution affect the polymerization rate, suggesting the participation of ionic interactions in the polymerization. Comparison of the molecular models of the S-crystallin and sigma-GST suggests that an extra ion-pair between Asp-101 and Arg-14 in S-crystallin contributes to stabilizing the protomer. Furthermore, the molecular surface of S-crystallin has a protruding Lys-208 on one side and a complementary patch of aspartate residues (Asp-90, Asp-94, Asp-101, Asp-102, Asp-179, and Asp-180) on the other side. We propose a molecular model for the S-crystallin polymer in vivo, which involves side-by-side associations of Lys-208 from one protomer and the aspartate patch from another protomer that allows the formation of a polymeric structure spontaneously into a liquid crystal structure in the lens.
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PMID:Molecular basis for the polymerization of octopus lens S-crystallin. 1073 85

It is established that the diverse, multifunctional crystallins are responsible for the optical properties of the cellular, transparent lens of the complex eyes of vertebrates and invertebrates. Lens crystallins often differ among species and may be enzymes or stress proteins. I present here the idea that abundant water-soluble enzymes and other proteins may also be used for cellular transparency in the epithelial cells and, possibly, stromal keratocytes of the cornea. Aldehyde dehydrogenases and transketolase are among the putative "corneal crystallins" in mammals, and gelsolin may be a corneal crystallin in the zebrafish. In invertebrates, the glutathione S-transferase-related S-crystallins of the lens appear to be used also as corneal crystallins in the squid, and an aldehyde dehydrogenase-related protein is the crystallin in the lens and, possibly, cornea of the scallop. The use of abundant, taxon-specific water-soluble proteins as crystallins for cellular transparency in the cornea would provide a new conceptual link between this tissue and the lens.
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PMID:Review: A case for corneal crystallins. 1080 28

It has been reported that alphaA-crystallin has greater protective effects against apoptosis in lens epithelial cells than alphaB-crystallin [Andley, Song, Wawrousek, Fleming and Bassnett (2000) J. Biol. Chem. 275, 36823-36831]. Because the alphaA-crystallin proteins are specifically expressed in the vertebrate lens, we examine the non-specific properties of both alphaA- and alphaB-crystallins in an Escherichia coli system. E. coli cells were transformed with the inducible protein expression vector pET-11a, harbouring the gene for either human alphaA- or alphaB-crystallin, and two other control plasmids, pET-1la vector alone or pGEX-2T vector encoding GST (glutathione S-transferase). These cells were exposed to various stress conditions, such as cold-shock at 4 degrees C or extremely low or high pH environments (pH 4.7 or pH 8.0) for 6 h, and survival of the host cells and the solubility of the expressed target proteins in the cytosol were examined. Under these stress conditions, the cells expressing alphaB-crystallin protein demonstrated significantly improved survival when compared with the other cells, and the expressed protein in the cytosol was almost soluble, in contrast with the alphaA-crystallin protein. Differences in the amino acid sequence between the proteins in a phenylalanine-rich region next to the N-terminal consensus alpha-crystallin domain was considered to be responsible for chaperone activity and cell survival.
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PMID:Differences in properties between human alphaA- and alphaB-crystallin proteins expressed in Escherichia coli cells in response to cold and extreme pH. 1282 11

The crystallins account for 80-90% of the water-soluble proteins of the transparent lens. These diverse proteins are responsible for the optical properties of the lens and have been recruited from metabolic enzymes and stress proteins. They often differ among species (i.e. are taxon-specific) and may be expressed outside of the lens where they have non-refractive roles (a situation we call gene sharing). Crystallin recruitment has occurred by changes in gene regulation resulting in high lens expression. Duck lactate dehydrogenase/epsilon-crystallin and alpha-enolase/tau-crystallin are each encoded in single-copy genes, consistent with these enzymes acquiring a crystallin role, without loss of their nonlens metabolic function, by a change in gene regulation in the absence of gene duplication. The small heat shock protein/alpha-crystallins and avian argininosuccinate lyase/delta-crystallins were also recruited by a change in gene regulation leading to high lens expression, except this was followed by a gene duplication with further lens specialization of the alphaA and the delta1 (in chickens) crystallin genes. Cephalopod (squid and octopus) S-crystallins were recruited from glutathione S-transferase apparently after duplication of the original gene encoding the enzyme, although this remains uncertain. We speculate that one of the new genes (glutathione S-transferase/S11-crystallin) specialized for lens expression by a change in gene regulation and subsequently duplicated many times to form the lens-specialized, multiple S-crystallins that lack enzymatic activity. That similar transcription factors (e.g. Pax-6, retinoic acid receptors, maf, Sox, AP-1, CREB) regulate different crystallin genes suggest that common features of lens-specific expression have played a pivotal role for recruiting the diverse, multifunctional proteins as crystallins.
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PMID:Crystallin genes: specialization by changes in gene regulation may precede gene duplication. 1283 92

Multiple sclerosis (MS) is a complex autoimmune disorder of the CNS with both genetic and environmental contributing factors. Clinical symptoms are broadly characterized by initial onset, and progressive debilitating neurological impairment. In this study, RNA from MS chronic active and MS acute lesions was extracted, and compared with patient matched normal white matter by fluorescent cDNA microarray hybridization analysis. This resulted in the identification of 139 genes that were differentially regulated in MS plaque tissue compared to normal tissue. Of these, 69 genes showed a common pattern of expression in the chronic active and acute plaque tissues investigated (Pvalue<0.0001, rho=0.73, by Spearman's rho analysis); while 70 transcripts were uniquely differentially expressed (> or = 1.5-fold) in either acute or chronic active tissues. These results included known markers of MS such as the myelin basic protein (MBP) and glutathione S-transferase (GST) M1, nerve growth factors, such as nerve injury-induced protein 1 (NINJ1), X-ray and excision DNA repair factors (XRCC9 and ERCC5) and X-linked genes such as the ribosomal protein, RPS4X. Primers were then designed for seven array-selected genes, including transferrin (TF), superoxide dismutase 1 (SOD1), glutathione peroxidase 1 (GPX1), GSTP1, crystallin, alpha-B (CRYAB), phosphomannomutase 1 (PMM1) and tubulin beta-5 (TBB5), and real time quantitative (Q)-PCR analysis was performed. The results of comparative Q-PCR analysis correlated significantly with those obtained by array analysis (r=0.75, Pvalue<0.01, by Pearson's bivariate correlation). Both chronic active and acute plaques shared the majority of factors identified suggesting that quantitative, rather than gross qualitative differences in gene expression pattern may define the progression from acute to chronic active plaques in MS.
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PMID:Quantitative and qualitative changes in gene expression patterns characterize the activity of plaques in multiple sclerosis. 1462 84

AlphaA- and alphaB-crystallins are distinct antiapoptotic regulators. Regarding the antiapoptotic mechanisms, we have recently demonstrated that alphaB-crystallin interacts with the procaspase-3 and partially processed procaspase-3 to repress caspase-3 activation. Here, we demonstrate that human alphaA- and alphaB-crystallins prevent staurosporine-induced apoptosis through interactions with members of the Bcl-2 family. Using GST pulldown assays and coimmunoprecipitations, we demonstrated that alpha-crystallins bind to Bax and Bcl-X(S) both in vitro and in vivo. Human alphaA- and alphaB-crystallins display similar affinity to both proapoptotic regulators, and so are true with their antiapoptotic ability tested in human lens epithelial cells, human retina pigment epithelial cells (ARPE-19) and rat embryonic myocardium cells (H9c2) under treatment of staurosporine, etoposide or sorbitol. Two prominent mutants, R116C in alphaA-crystallin and R120G, in alphaB-crystallin display much weaker affinity to Bax and Bcl-X(S). Through the interaction, alpha-crystallins prevent the translocation of Bax and Bcl-X(S) from cytosol into mitochondria during staurosporine-induced apoptosis. As a result, alpha-crystallins preserve the integrity of mitochondria, restrict release of cytochrome c, repress activation of caspase-3 and block degradation of PARP. Thus, our results demonstrate a novel antiapoptotic mechanism for alpha-crystallins.
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PMID:Human alphaA- and alphaB-crystallins bind to Bax and Bcl-X(S) to sequester their translocation during staurosporine-induced apoptosis. 1475 12

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


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