Glutathione S-transferases: an overview in cancer research (original) (raw)
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Scientific Reports, 2013
The gene for glutathione-S-transferase (GST) M1 (GSTM1), a member of the GST-superfamily, is widely studied in cancer risk with regard to the homozygous deletion of the gene (GSTM1 null), leading to a lack of corresponding enzymatic activity. Many of these studies have reported inconsistent findings regarding its association with cancer risk. Therefore, we employed in silico, in vitro, and in vivo approaches to investigate whether the absence of a functional GSTM1 enzyme in a null variant can be compensated for by other family members. Through the in silico approach, we identified maximum structural homology between GSTM1 and GSTM2. Total plasma GST enzymatic activity was similar in recruited individuals, irrespective of their GSTM1 genotype (positive/null). Furthermore, expression profiling using real-time PCR, western blotting, and GSTM2 overexpression following transient knockdown of GSTM1 in HeLa cells confirmed that the absence of GSTM1 activity can be compensated for by the overexpression of GSTM2. G lutathione-S-transferases (GSTs) belong to a superfamily of ubiquitous, multifunctional dimeric cytosolic enzymes that play a very important role in the Phase II detoxification (or biotransformation) pathway in humans and confer protection against a wide array of toxic insults 1,2. Several GST isoforms have been identified and characterised, forming seven distinct classes: a, m, p, o, t, k, and f 3,4. Functionally, most GSTs catalyse the conjugation of the nucleophilic tripeptide glutathione to a wide range of electrophilic substrates for detoxification. However, the conjugation reaction can occasionally lead to the formation of compounds that are far more toxic than the initial substrate, thereby leading to disease outcomes 1,5,6. Interestingly, a null variant is encountered for two members, GSTT1 and GSTM1, whereby the entire gene is homozygously deleted in a considerable proportion of different populations, resulting in the complete absence of the corresponding enzyme activity 7,8. The GSTM1 gene is highly polymorphic and is located on chromosome 1p13.3. A wide range of variation in GSTM1 homozygous deletion polymorphism (approximately 20-67%) has been observed globally with regard to various ethnicities 9-12. It is often hypothesised that, due to the lack of functional GSTT1 and/or GSTM1, the null phenotype is unable to efficiently perform the conjugation reaction (biotransformation) and the subsequent elimination of toxic products via urine and bile. The null variant of GSTM1 is of particular interest, as a plethora of studies have demonstrated the difference in susceptibility, exposure to environmental toxicants, resistance to chemotherapy treatment, variability in drug response, manifestation of several diseases, and, most importantly, cancerous outcomes. The four other members of the GSTm subfamily, i.e., GSTM2, GSTM3, GSTM4, and GSTM5, exhibit high levels of sequence homology and substrate specificity with GSTM1 13. Among these genes, GSTM1 has largely been studied due to its null genotype. Although a large number of studies have attempted to associate the GSTM1-null genotype with cancer risk, the results are inconclusive. Several studies have attempted to identify the association of GSTM1 null with cancer risk through meta-analysis using the existing literature; however, these analyses failed to show a significant association of GSTM1 with cancer 14-20. These observations prompted us to search for the functional relevance of this ''well known gene'' with other family members that are relatively less studied. A possible explanation of the apparently inconsistent results could be that other members of the GST family compensate for the absence of a functional GSTM1 enzyme. In this study, we attempted to ascertain whether
GLUTATHIONE S-TRANSFERASES: A BRIEF ON CLASSIFICATION AND GSTM1-T1 ACTIVITY
The glutathione S-transferase (GST) isoenzyme superfamilies detoxify a wide-range of toxic chemicals and environmental substances are extensively expressed in mammalian tissues. Liver and pancreas are the sites where cytosolic Phase I and phase II biotransformation GSTs enzymes have characteristic expression. GSTs play a key role in the deactivation of reactive oxygen species (ROS) and the metabolism of lipids, chemotherapeutic agents. GSTs are mainly involved in conjugation of reduced glutathione (GSH) with diverse substrates specificity and it is possible that genetic variations in these enzymes will influence cellular response to the environmental agents. GSTs are overexpressed in response to a chemical or oxidative stress as an adaptive physiology and upregulated in cancerous state of organ or tissue. GSTs are essentially involved in susceptibility to various forms of cancer as they are vital in detoxification mechanism to metabolize the environmental carcinogens. GSTM1 encodes for a class mu GST isoenzyme involved in polycyclic aromatic hydrocarbons (PAHs) detoxification. The substrates of GSTM1 include benzo(a)pyrene, benzo(c)phenanthrene, benzo(g)chrysene and other carcinogens. They can catalyze in-vitro GSH conjugation with several potent carcinogenic epoxides including aflatoxin B1(AFB1)8,9-epoxide and electrophilic metabolites of PAHs present in tobacco smoke. Ethylene dibromide, p-nitrobenzyl chloride, p-nitrophenetyl bromide, methyl chloride, and methyl iodide, are known substrates for GSTT1 or GST Theta (θ). GST Theta is most primitive among other known GSTs and widely expressed in nature.
Glutathione S-transferases, structure, regulation, and therapeutic implications
Journal of Biological Chemistry, 1993
The glutathione S-transferases (GSTs)' are a family of enzymes that catalyze the nucleophilic addition of the thiol of reduced glutathione to a variety of electrophiles (1-9). In addition, the GSTs bind with varying affinities a variety of hydrophobic compounds such as heme, bilirubin, polycyclic aromatic hydrocarbons, and dexamethasone (1-9). It is now generally accepted that the GSTs are encoded by at least five different gene families (9-12). Four of the gene families encode the cytosolic GSTs whereas the fifth encodes a microsomal form of the enzyme (13, 14). In this review, we have focused on three GST research areas: 1) structure-function analysis of GSTs; 2) regulation of GST expression; and 3) GSTs as therapeutic targets in disease. Since our review will not be comprehensive, we would like to direct the reader to several excellent recent reviews focusing on various aspects of GSTs (4,8,9).
We report the isolation of three full-length cDNAs corresponding to the mRNAs of closely related glutathione S-transferase (GST) Pi genes, designated hGSTP1*A, hGSTP1*B, and hGSTP1*C, expressed in normal cells and malignant gliomas. The variant cDNAs result from A 3 G and C 3 T transitions at nucleotides 313 and 341, respectively. The transitions changed codon 104 from ATC (Ile) in hGSTP1*A to GTC (Val) in hGSTP1*B and hGSTP1*C and changed codon 113 from GCG (Ala) to GTG (Val) in hGSTP1*C. Both amino changes are in the electrophile-binding active site of the GST Pi peptide. Computer modeling of the deduced crystal structures of the encoded peptides showed significant deviations in the interatomic distances of critical electrophile-binding active site amino acids as a consequence of the amino acid changes. The encoded proteins expressed in Escherichia coli and purified by GSH affinity chromatography showed a 3-fold lower K m (CDNB) and a 3– 4-fold higher K cat /K m for the hGSTP1*A encoded protein than the proteins encoded by hGSTP1*B and hGSTP1*C. Analysis of 75 cases showed the relative frequency of hGSTP1*C to be 4-fold higher in malignant gliomas than in normal tissues. These data provide conclusive molecular evidence of allelopolymorphism of the human GST Pi gene locus, resulting in active, functionally different GST Pi proteins, and should facilitate studies of the role of this gene in xenobiotic metabolism, cancer, and other human diseases.
Glutathione S-transferase Activity in Diagnostic Pathology
Journal of Postgenomics Drug & Biomarker Development, 2015
etc. were used to collate relevant articles. The results were then crossreferenced to generate a total number of 125 references cited in this review. Functions of glutathione S-transferases The functions of GSTs have been classified into two general categories [19,20]. As intracellular binding proteins [2,21,22], GSTs function on a broad scale in solubilizing and transport of substances much as the extracellular functions of albumin described elsewhere [23,24]. The GST from rat liver, designated as transferase B, has been shown to be identical to the bilirubin binding protein or 'ligandins' [25]. Although ligandins have high affinity for endogenous compounds such as bile acids, haemin, bilirubin, fatty acids and steroids [16,18,22], whose conjugates are eventually sequestered [26], the bound GSTs are devoid of catalytic processing and do not form glutathione conjugates with their substrates [18,27]. Another specific protective role of GST as ligandin is the specific binding of intra-erythrocyte GSTP1-1 isoform to Jun-kinase, a pro-apoptotic enzyme that becomes inactive when bound to GST [26,28]. The second major function is the protection of cellular components [29,30] by the preferential reaction of electrophilic agents with GSH through the enzymatic action of GSTs, and thereby prevents the reaction of electrophiles with cellular nucleophiles. The enzyme may also detoxify certain extremely reactive substances by direct covalent binding to electrophilic agents [1,22,31]. For the most part, GSTs catalyze the conjugation of electrophilic groups of hydrophobic drugs and xenobiotics to form glutathione-thioethers [32]. These thioethers are converted to mercapturic acid by the sequential actions of γ-glutamyl transpeptidase, depeptidase and N-acetylase [2,15,33] prior to the eventual elimination of the hydrophilic conjugates. Reactive oxygen and nitrogen species (ROS/RNS) can alter the
Archives of Biochemistry and Biophysics, 1999
In order to elucidate the protective role of glutathione S-transferases (GSTs) against oxidative stress, we have investigated the kinetic properties of the human ␣-class GSTs, hGSTA1-1 and hGSTA2-2, toward physiologically relevant hydroperoxides and have studied the role of these enzymes in glutathione (GSH)-dependent reduction of these hydroperoxides in human liver. We have cloned hGSTA1-1 and hGSTA2-2 from a human lung cDNA library and expressed both in Escherichia coli. Both isozymes had remarkably high peroxidase activity toward fatty acid hydroperoxides, phospholipid hydroperoxides, and cumene hydroperoxide. In general, the activity of hGSTA2-2 was higher than that of hGSTA1-1 toward these substrates. For example, the catalytic efficiency (k cat /K m) of hGSTA1-1 for phosphatidylcholine (PC) hydroperoxide and phosphatidylethanolamine (PE) hydroperoxide was found to be 181.3 and 199.6 s ؊1 mM ؊1 , respectively, while the catalytic efficiency of hGSTA2-2 for PC-hydroperoxide and PE-hydroperoxide was 317.5 and 353 s ؊1 mM ؊1 , respectively. Immunotitration studies with human liver extracts showed that the antibodies against human ␣-class GSTs immunoprecipitated about 55 and 75% of glutathione peroxidase (GPx) activity of human liver toward PC-hydroperoxide and cumene hydroperoxide, respectively. GPx activity was not immunoprecipitated by the same antibodies from human erythrocyte hemolysates. These results show that the ␣-class GSTs contribute a major portion of GPx activity toward lipid hydroperoxides in human liver. Our results also suggest that GSTs may be involved in the reduction of 5-hydroperoxyeicosatetrae-noic acid, an important intermediate in the 5-lipoxygenase pathway.
Glutathione-S-Transferases: As Signaling Molecules
The Glutathione-s-transferases exist as cytosolic, mitochondrial, and microsomal can participate in signal transduction by not phosphorylating any factor that is directly involved in cell growth and death. This signal transduction is considered to be a new way of implication in cell metabolic pathways due to the influence of external, such as xenobiotics and UV radiation, and internal, such as oxidative stress, free radicals, etc., agents. The GST binding assay studies revealed that they participate in the inhibition of various proteins, for example, phosphoproteins, AP-1, JNK, etc., in the systems to regulate cell mechanisms during cell synthesis.
FEBS Journal, 2011
Glutathione (GSH) conjugating enzymes, glutathione S-transferases (GSTs) are present in different subcellular compartments including cytosol, mitochondria, endoplasmic reticulum, nucleus and plasma membrane. The regulation and function of GSTs have implications in cell growth, oxidative stress, as well as in disease progression and prevention. Of the several mitochondria localized forms, GSTK (GST kappa) is mitochondria-specific since it contains Nterminal canonical and cleavable mitochondria targeting signal. Other forms, like GST alpha, mu and pi purified from mitochondria are similar to the cytosolic molecular forms or "echoproteins". Altered GST expression has been implicated in hepatic, cardiac and neurological diseases. Mitochondria-specific GSTK has also been implicated in obesity, diabetes and related metabolic disorders. Studies have shown that silencing the GSTA4 (GST alpha) gene resulted in mitochondrial dysfunction, as was also seen in GSTA4 null mice which could contribute to insulin resistance in type 2 diabetes. This review highlights the significance of mitochondrial GST pool, particularly the mechanism and significance of dual targeting of GSTA4-4 under in vitro and in vivo conditions. GSTA4-4 is targeted in the mitochondria by activation of the internal cryptic signal present at the C-terminus of the protein by protein kinase-dependent phosphorylation and cytosolic heat shock protein (Hsp70) chaperon. Mitochondrial GSTpi, on the other hand, has been shown to have two uncleaved cryptic signals rich in positively charged amino acids at the Nterminal region. Both physiological and pathophysiological implications of GST translocation to mitochondria have been discussed in this review.
Archives of Biochemistry and Biophysics, 1991
Human muscle glutathione S-transferase isozyme, GST{ (~15.2) has been purified by three different methods using immunoaffinity chromatography, DEAE cellulose chromatography, and isoelectric focusing. GST{ prepared by any of the three methods does not recognize antibodies raised against the a, ~1, or x class glutathione S-transferases of human tissues. GSTf has a blocked Nterminus and its peptide fingerprints also indicate it to be distinct from the a, N, or u class isozymes. As compared to GSTs of a, cc, and x classes, GSTt displays higher activities toward t-stilbene oxide and Leukotriene A4 methyl ester. GST{ also expresses GSH-peroxidase activity toward hydrogen peroxide. The K,s of GST{ for CDNB and GSH were comparable to those reported for other human GSTs but its V,, for CDNB, 7620 mol/mol/ min, was found to be considerably higher than that reported for other human GSTs. The kinetics of inhibition of GST{ by hematin, bile acids, and other inhibitors also indicate that it was distinct from the three classes of GST isozymes. These studies suggest that GST{ corresponds to a locus distinct from GSTl, GST2, and GST3 and probably corresponds to the GST4 locus as suggested previously by Laisney et al. (1984, Human Cenet. 68, 22 l-227). The results of peptide fingerprints and kinetic analysis indicate that as compared to the a and a class isozymes, GST{ has more structural and functional similarities with the p class isozymes. Besides GSTC several