Glutathione s-transferase

Details Glutathione s-transferase Glutathione s-transferase

1999-05-11

2001-06-19

Graser, Jennifer (Department: 1641)

Drug, bio-affecting and body treating compositions

Enzyme or coenzyme containing

Transferases

C424S009340, C424S133100, C424S134100, C424S139100, C424S200100, C424S809000, C435S007700, C435S015000, C435S097000, C435S193000, C530S387300, C530S866000

Reexamination Certificate

active

06248325

ABSTRACT:

FIELD OF THE INVENTION
This invention relates to nucleic acid and amino acid sequences of a novel glutathione s-transferase and to the use of these sequences in the diagnosis, prevention, and treatment of cancer.
BACKGROUND OF THE INVENTION
The glutathione s-transferases (GST) are a ubiquitous family of enzymes with dual substrate specificities that perform important biochemical functions of xenobiotic biotransformation and detoxification, drug metabolism, and protection of tissues against peroxidative damage. The basic reaction catalyzed by these enzymes is the conjugation of an electrophile with reduced glutathione (GSH), results in either activation or deactivation/detoxification of the chemical. The absolute requirement for binding reduced GSH to a wide variety of chemicals necessitates a diversity in GST structures in various organisms and cell types.
GSTs are homodimeric or heterodimeric proteins localized in the cell cytosol. The major isozymes share common structural and catalytic properties and in man have been classified into four major classes, Alpha, Mu, Pi, and Theta. The two largest classes, Alpha and Mu, are identified by their respective protein isoelectric point, pI ~7.5-9.0 Alpha, and pI ~6.6 (Mu). Each GST possesses a common binding site for GSH and a variable hydrophobic binding site. The hydrophobic binding site in each isozyme is specific for particular electrophilic substrates. Specific amino acid residues within GSTs have been identified as important for these binding sites and for catalytic activity. Residues Q67, T68, D101, E104, and R131 are important for the binding of GSH (Lee, H-C et al. (1995) J. Biol. Chem. 270: 99-109). Residues R13, R20, and R69 are important for the catalytic activity of GST. (Stenberg G et al. (1991) Biochem. J. 274: 549-55).
In most cases, GSTs perform the beneficial function of deactivation and detoxification of potentially mutagenic and carcinogenic chemicals. However, in some cases their action is detrimental and results in activation with consequent mutagenic and carcinogenic effects. Some forms of rat and human GSTs are reliable preneoplastic markers that aid in the detection of carcinogenesis. Expression of human GSTs in bacterial strains, such as Salmonella typhimurium, used in the well known Ames test for mutagenicity, has helped to establish the role of these enzymes in mutagenesis. Dihalomethanes, which produce liver tumors in mice, are believed to be activated by GST. This view is supported by the finding that dihalomethanes are more mutagenic in bacterial cells expressing human GST than in untransfected cells (Thier, R. et al. (1993) Proc. Natl. Acad. Sci. USA 90: 8567-80). The mutagenicity of ethylene dibromide and ethylene dichloride is increased in bacterial cells expressing the human Alpha GST, A1-1, while the mutagenicity of aflatoxin B1 is substantially reduced by enhancing the expression of GST (Simula, T. P. et al. (1993) Carcinogenesis 14: 1371-6). Thus, control of GST activity may be useful in the control of mutagenesis and carcinogenesis.
GST has been implicated in the acquired resistance of many cancers to drug treatment, the phenomenon known as multi-drug resistance (MDR). MDR occurs when a cancer patient is treated with a cytotoxic drug such as cyclophosphamide and subsequently becomes resistant to this drug and to a variety of other cytotoxic agents as well. Increased GST levels are associated with some of these drug resistant cancers, and it is believed that this increase occurs in response to the drug agent which is deactivated by the GST catalyzed GSH conjugation reaction. The increased GST levels protect the cancer cells from other cytotoxic agents for which GST has affinity. Increased levels of A1-1 in tumors have been linked to drug resistance induced by cyclophosphamide treatment (Dirven H. A. et al. (1994) Cancer Res. 54: 6215-20). Thus, control of GST activity in cancerous tissues may be useful in treating MDR in cancer patients.
The discovery of polynucleotides encoding novel glutathione s-transferases and the molecules themselves, provide the means to further investigate the role of these enzymes in chemical mutagenesis and carcinogenesis and in drug resistance and cancer. In particular, the ability to control GST activity may be useful in the prevention of carcinogenesis where the risk of exposure to carcinogens or mutagens is high and in the prevention and treatment of MDR in cancerous tissues.
SUMMARY OF THE INVENTION
The present invention features a novel human glutathione s-transferase hereinafter designated HGST and characterized as having similarity to Alpha class GSTs from normal human liver pGTH2; GI 825605) human hepatoma, (A1-1; GI 259141), and mouse lung, (GST 5.7; GI 193710). Accordingly, the invention features a substantially purified HGST having the amino acid sequence, SEQ ID NO:1.
One aspect of the invention features isolated and substantially purified polynucleotides that encode HGST. In a particular aspect, the polynucleotide is the nucleotide sequence of SEQ ID NO:2.
The invention also relates to a polynucleotide sequence comprising the complement of SEQ ID NO:2 or variants thereof. In addition, the invention features polynucleotide sequences which hybridize under stringent conditions to SEQ ID NO:2.
The invention additionally features nucleic acid sequences encoding polypeptides, oligonucleotides, peptide nucleic acids (PNA), fragments, portions or antisense molecules thereof, and expression vectors and host cells comprising polynucleotides that encode HGST. The present invention also features antibodies which bind specifically to HGST, and pharmaceutical compositions comprising substantially purified HGST. The invention also features the use of agonists and antagonists of HGST.


REFERENCES:
patent: 5658727 (1997-08-01), Barbas et al.
patent: WO 96 02674 (1996-02-01), None
Dirven et al. 1994. cancer research. 54:6215-20.*
Liu et al. 1993. Biochimica et Biophysica Acta. 1216:332-334.*
Orlandi et al. 1989. PNAS. 86:3833-3837.*
Stenberg et al. Biochem J. 284:313-319, 1992.*
Stenberg et al. 1991. Biochem J. 274:549-555.*
Stenberg et al. 1992. Protein Expr Purif. 3:80-84.*
Zimniak et al. 1992. FEBS. 313(2): 173-176.*
Lee, HC et al., “A molecular genetic approach for the identification of essential residues in human glutathione S-transferse function inEscherichia coli”J. Biol. Chem.270(1):99-109 (1995).
Stenberg, G et al., “Effects of directed mutagenesis on conserved arginine residues in a human Class Alpha gluthathione transferase”Biochem. J.274:549-555 (1991).
Thier, R et al., “Expression of mammalian glutathione S-transferase 5—5 inSalmonella typhimuriumTA1535 leads to base-pair mutations upon exposure to dihalomethanes”Proc. Natl. Acad. Sci. USA90:8567-8580 (1993).
Simula, TP et al., Human glutathione S-transferae-expressingSalmonella typhimuriumtester strains to study the activation/detoxification of mutagenic compounds: studies with halogenated compounds, aromatic amines and aflatoxin B1Carcinogenesis14(7):1371-1376 (1993.
Dirven, Haam et al., “Involvement of Human Glutathione S-Transferase Isoenzymes in the Conjugation of Cyclophosphamide Metabolites with Glutathione”Cancer Res.54:6215-6220 (1994).
Klöne, A et al., “Cloning, sequencing and characterization of the human Alpha glutathione S-transferase gene corresponding to the cDNA clone pGTH2”Biochem. J.285:925-928 (1992) (Accession GI 825605).
Stenberg, G et al., “Heterologous expression of recombinant human glutathione transferase A1-1 from a hepatoma cell line”Protein Expr Purif3:80-84 (1992) (Accession GI 259141).
Zimniak, P et al., “A subgroup of class &agr; glutathione S-transferases Cloning of cDNA for mouse lung glutathione S-transferase GST 5.7”FEBS Lett313(2):173-176 (1992) (Accession GI 193710).
Database: EMBL Sequences, EMBL, Heidelberg, FRG Accession No. H27975, Jul. 18, 1995, Hillier L. Et al., “H.sapienscDNA clone 162882 similar to mouse glutathione S-transferase GST5.7”, XP002057353.
Database: EMBL Sequences, EMBL, Heidelberg, FRG Accession No. WO5487, May 8, 1996, Hillier, L., et al., “Soares fe

Affiliated with

Also associated with

Rating

Glutathione s-transferase does not yet have a rating. At this time, there are no reviews or comments for this patent.

If you have personal experience with Glutathione s-transferase, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Glutathione s-transferase will most certainly appreciate the feedback.

Rate now

Comments { 0 }

Profile ID: LFUS-PAI-O-2503331