Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 2002 Aug 1;365(Pt 3):685–691. doi: 10.1042/BJ20020127

Mu-class glutathione transferase from Xenopus laevis: molecular cloning, expression and site-directed mutagenesis.

Antonella De Luca 1, Bartolo Favaloro 1, Stefania Angelucci 1, Paolo Sacchetta 1, Carmine Di Ilio 1
PMCID: PMC1222732  PMID: 11991804

Abstract

A cDNA encoding a Mu-class glutathione transferase (XlGSTM1-1) has been isolated from a Xenopus laevis liver library, and its nucleotide sequence has been determined. XlGSTM1-1 is composed of 219 amino acid residues with a calculated molecular mass of 25359 Da. Unlike many mammalian Mu-class GSTs, XlGSTM1-1 has a narrow spectrum of substrate specificity and it is also less effective in conjugating 1-chloro-2,4-dinitrobenzene. A notable structural feature of XlGSTM1-1 is the presence of the Cys-139 residue in place of the Glu-139, as well as the absence of the Cys-114 residue, present in other Mu-class GSTs, which is replaced by Ala. Site-directed mutagenesis experiments indicate that Cys-139 is not involved in the catalytic mechanism of XlGSTM1-1 but may be in part responsible for its structural instability, and experiments in vivo confirmed the role of this residue in stability. Evidence indicating that Arg-107 is essential for the 1-chloro-2,4-dinitrobenzene conjugation capacity of XlGSTM1-1 is also presented.

Full Text

The Full Text of this article is available as a PDF (339.6 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aceto A., Dragani B., Sacchetta P., Bucciarelli T., Angelucci S., Miranda M., Poma A., Amicarelli F., Federici G., di Ilio C. Developmental aspects of Bufo bufo embryo glutathione transferases. Mech Ageing Dev. 1993 May;68(1-3):59–70. doi: 10.1016/0047-6374(93)90140-m. [DOI] [PubMed] [Google Scholar]
  2. Amicarelli F., Ragnelli A. M., Aimola P., Cattani F., Bonfigli A., Zarivi O., Miranda M., Di Ilio C. Developmental expression and distribution of amphibian glutathione transferases. Biochim Biophys Acta. 2001 Apr 3;1526(1):77–85. doi: 10.1016/s0304-4165(01)00103-9. [DOI] [PubMed] [Google Scholar]
  3. Angelucci Stefania, Sacchetta Paolo, De Luca Antonella, Moio Pasquale, Amicarelli Fernanda, Di Ilio Carmine. Glutathione transferase isoenzymes from frog (Xenopus laevis) liver and embryo. Biochim Biophys Acta. 2002 Jan 15;1569(1-3):81–85. doi: 10.1016/s0304-4165(01)00238-0. [DOI] [PubMed] [Google Scholar]
  4. Armstrong R. N. Structure, catalytic mechanism, and evolution of the glutathione transferases. Chem Res Toxicol. 1997 Jan;10(1):2–18. doi: 10.1021/tx960072x. [DOI] [PubMed] [Google Scholar]
  5. Board P. G., Baker R. T., Chelvanayagam G., Jermiin L. S. Zeta, a novel class of glutathione transferases in a range of species from plants to humans. Biochem J. 1997 Dec 15;328(Pt 3):929–935. doi: 10.1042/bj3280929. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Board P. G., Coggan M., Chelvanayagam G., Easteal S., Jermiin L. S., Schulte G. K., Danley D. E., Hoth L. R., Griffor M. C., Kamath A. V. Identification, characterization, and crystal structure of the Omega class glutathione transferases. J Biol Chem. 2000 Aug 11;275(32):24798–24806. doi: 10.1074/jbc.M001706200. [DOI] [PubMed] [Google Scholar]
  7. Bochner B. R., Lee P. C., Wilson S. W., Cutler C. W., Ames B. N. AppppA and related adenylylated nucleotides are synthesized as a consequence of oxidation stress. Cell. 1984 May;37(1):225–232. doi: 10.1016/0092-8674(84)90318-0. [DOI] [PubMed] [Google Scholar]
  8. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1016/0003-2697(76)90527-3. [DOI] [PubMed] [Google Scholar]
  9. Bucciarelli T., Sacchetta P., Pennelli A., Cornelio L., Romagnoli R., Melino S., Petruzzelli R., Di Ilio C. Characterization of toad liver glutathione transferase. Biochim Biophys Acta. 1999 Apr 12;1431(1):189–198. doi: 10.1016/s0167-4838(99)00036-9. [DOI] [PubMed] [Google Scholar]
  10. Chen W. L., Hsieh J. C., Hong J. L., Tsai S. P., Tam M. F. Site-directed mutagenesis and chemical modification of cysteine residues of rat glutathione S-transferase 3-3. Biochem J. 1992 Aug 15;286(Pt 1):205–210. doi: 10.1042/bj2860205. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Cho S. G., Lee Y. H., Park H. S., Ryoo K., Kang K. W., Park J., Eom S. J., Kim M. J., Chang T. S., Choi S. Y. Glutathione S-transferase mu modulates the stress-activated signals by suppressing apoptosis signal-regulating kinase 1. J Biol Chem. 2001 Jan 18;276(16):12749–12755. doi: 10.1074/jbc.M005561200. [DOI] [PubMed] [Google Scholar]
  12. Di Ilio C., Sacchetta P., Lo Bello M., Caccuri A. M., Federici G. Selenium independent glutathione peroxidase activity associated with cationic forms of glutathione transferase in human heart. J Mol Cell Cardiol. 1986 Sep;18(9):983–991. doi: 10.1016/s0022-2828(86)80012-8. [DOI] [PubMed] [Google Scholar]
  13. ELLMAN G. L. Tissue sulfhydryl groups. Arch Biochem Biophys. 1959 May;82(1):70–77. doi: 10.1016/0003-9861(59)90090-6. [DOI] [PubMed] [Google Scholar]
  14. Habig W. H., Jakoby W. B. Assays for differentiation of glutathione S-transferases. Methods Enzymol. 1981;77:398–405. doi: 10.1016/s0076-6879(81)77053-8. [DOI] [PubMed] [Google Scholar]
  15. Hayes J. D., Pulford D. J. The glutathione S-transferase supergene family: regulation of GST and the contribution of the isoenzymes to cancer chemoprotection and drug resistance. Crit Rev Biochem Mol Biol. 1995;30(6):445–600. doi: 10.3109/10409239509083491. [DOI] [PubMed] [Google Scholar]
  16. Hayes J. D., Strange R. C. Potential contribution of the glutathione S-transferase supergene family to resistance to oxidative stress. Free Radic Res. 1995 Mar;22(3):193–207. doi: 10.3109/10715769509147539. [DOI] [PubMed] [Google Scholar]
  17. Jakobsson P. J., Morgenstern R., Mancini J., Ford-Hutchinson A., Persson B. Common structural features of MAPEG -- a widespread superfamily of membrane associated proteins with highly divergent functions in eicosanoid and glutathione metabolism. Protein Sci. 1999 Mar;8(3):689–692. doi: 10.1110/ps.8.3.689. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Ji X., Johnson W. W., Sesay M. A., Dickert L., Prasad S. M., Ammon H. L., Armstrong R. N., Gilliland G. L. Structure and function of the xenobiotic substrate binding site of a glutathione S-transferase as revealed by X-ray crystallographic analysis of product complexes with the diastereomers of 9-(S-glutathionyl)-10-hydroxy-9,10-dihydrophenanthrene. Biochemistry. 1994 Feb 8;33(5):1043–1052. doi: 10.1021/bi00171a002. [DOI] [PubMed] [Google Scholar]
  19. Ji X., Zhang P., Armstrong R. N., Gilliland G. L. The three-dimensional structure of a glutathione S-transferase from the mu gene class. Structural analysis of the binary complex of isoenzyme 3-3 and glutathione at 2.2-A resolution. Biochemistry. 1992 Oct 27;31(42):10169–10184. doi: 10.1021/bi00157a004. [DOI] [PubMed] [Google Scholar]
  20. Lardelli M., Lendahl U. Generating bacteriophage lambda sublibraries enriched for rare clones. Biotechniques. 1994 Mar;16(3):420–422. [PubMed] [Google Scholar]
  21. Lee H. C., Toung Y. P., Tu Y. S., Tu C. P. A molecular genetic approach for the identification of essential residues in human glutathione S-transferase function in Escherichia coli. J Biol Chem. 1995 Jan 6;270(1):99–109. doi: 10.1074/jbc.270.1.99. [DOI] [PubMed] [Google Scholar]
  22. Mannervik B., Alin P., Guthenberg C., Jensson H., Tahir M. K., Warholm M., Jörnvall H. Identification of three classes of cytosolic glutathione transferase common to several mammalian species: correlation between structural data and enzymatic properties. Proc Natl Acad Sci U S A. 1985 Nov;82(21):7202–7206. doi: 10.1073/pnas.82.21.7202. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Mannervik B., Awasthi Y. C., Board P. G., Hayes J. D., Di Ilio C., Ketterer B., Listowsky I., Morgenstern R., Muramatsu M., Pearson W. R. Nomenclature for human glutathione transferases. Biochem J. 1992 Feb 15;282(Pt 1):305–306. doi: 10.1042/bj2820305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Mannervik B., Danielson U. H. Glutathione transferases--structure and catalytic activity. CRC Crit Rev Biochem. 1988;23(3):283–337. doi: 10.3109/10409238809088226. [DOI] [PubMed] [Google Scholar]
  25. Meyer D. J., Coles B., Pemble S. E., Gilmore K. S., Fraser G. M., Ketterer B. Theta, a new class of glutathione transferases purified from rat and man. Biochem J. 1991 Mar 1;274(Pt 2):409–414. doi: 10.1042/bj2740409. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Patskovsky Y. V., Patskovska L. N., Listowsky I. Functions of His107 in the catalytic mechanism of human glutathione S-transferase hGSTM1a-1a. Biochemistry. 1999 Jan 26;38(4):1193–1202. doi: 10.1021/bi982164m. [DOI] [PubMed] [Google Scholar]
  27. Patskovsky Y. V., Patskovska L. N., Listowsky I. The enhanced affinity for thiolate anion and activation of enzyme-bound glutathione is governed by an arginine residue of human Mu class glutathione S-transferases. J Biol Chem. 2000 Feb 4;275(5):3296–3304. doi: 10.1074/jbc.275.5.3296. [DOI] [PubMed] [Google Scholar]
  28. Pemble S. E., Wardle A. F., Taylor J. B. Glutathione S-transferase class Kappa: characterization by the cloning of rat mitochondrial GST and identification of a human homologue. Biochem J. 1996 Nov 1;319(Pt 3):749–754. doi: 10.1042/bj3190749. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Raghunathan S., Chandross R. J., Kretsinger R. H., Allison T. J., Penington C. J., Rule G. S. Crystal structure of human class mu glutathione transferase GSTM2-2. Effects of lattice packing on conformational heterogeneity. J Mol Biol. 1994 May 20;238(5):815–832. doi: 10.1006/jmbi.1994.1336. [DOI] [PubMed] [Google Scholar]
  30. Rowe J. D., Patskovsky Y. V., Patskovska L. N., Novikova E., Listowsky I. Rationale for reclassification of a distinctive subdivision of mammalian class Mu glutathione S-transferases that are primarily expressed in testis. J Biol Chem. 1998 Apr 17;273(16):9593–9601. doi: 10.1074/jbc.273.16.9593. [DOI] [PubMed] [Google Scholar]
  31. Sacchetta P., Petruzzelli R., Melino S., Bucciarelli T., Pennelli A., Amicarelli F., Miranda M., Di Ilio C. Amphibian embryo glutathione transferase: amino acid sequence and structural properties. Biochem J. 1997 Mar 1;322(Pt 2):679–680. doi: 10.1042/bj3220679. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Shan S., Armstrong R. N. Rational reconstruction of the active site of a class mu glutathione S-transferase. J Biol Chem. 1994 Dec 23;269(51):32373–32379. [PubMed] [Google Scholar]
  33. Simons P. C., Vander Jagt D. L. Purification of glutathione S-transferases from human liver by glutathione-affinity chromatography. Anal Biochem. 1977 Oct;82(2):334–341. doi: 10.1016/0003-2697(77)90169-5. [DOI] [PubMed] [Google Scholar]
  34. Stanley J. S., Benson A. M. The conjugation of 4-nitroquinoline 1-oxide, a potent carcinogen, by mammalian glutathione transferases. 4-Nitroquinoline 1-oxide conjugation by human, rat and mouse liver cytosols, extrahepatic organs of mice and purified mouse glutathione transferase isoenzymes. Biochem J. 1988 Nov 15;256(1):303–306. doi: 10.1042/bj2560303. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES