title: Arabidopsis Glutathione Transferases U24 and U25 Exhibit a Range of Detoxification Activities with the Environmental Pollutant and Explosive, 2,4,6-Trinitrotoluene.
abstract: The explosive 2,4,6-trinitrotoluene (TNT) is a major worldwide military pollutant. The presence of this toxic and highly persistent pollutant, particularly at military sites and former manufacturing facilities, presents various health and environmental concerns. Due to the chemically resistant structure of TNT, it has proven to be highly recalcitrant to biodegradation in the environment. Here, we demonstrate the importance of two glutathione transferases (GSTs), GST-U24 and GST-U25, from Arabidopsis (Arabidopsis thaliana) that are specifically up-regulated in response to TNT exposure. To assess the role of GST-U24 and GST-U25, we purified and characterized recombinant forms of both enzymes and demonstrated the formation of three TNT glutathionyl products. Importantly, GST-U25 catalyzed the denitration of TNT to form 2-glutathionyl-4,6-dinitrotoluene, a product that is likely to be more amenable to subsequent biodegradation in the environment. Despite the presence of this biochemical detoxification pathway in plants, physiological concentrations of GST-U24 and GST-U25 result in only a limited innate ability to cope with the levels of TNT found at contaminated sites. We demonstrate that Arabidopsis plants overexpressing GST-U24 and GST-U25 exhibit significantly enhanced ability to withstand and detoxify TNT, properties that could be applied for in planta detoxification of TNT in the field. The overexpressing lines removed significantly more TNT from soil and exhibited a corresponding reduction in glutathione levels when compared with wild-type plants. However, in the absence of TNT, overexpression of these GSTs reduces root and shoot biomass, and although glutathione levels are not affected, this effect has implications for xenobiotic detoxification.
authors.affiliation: Centre for Novel Agricultural Products (V.G., K.T., H.S., E.J.J., E.L.R., N.C.B.) and Department of Biology (A.S.B., J.R.P.), University of York, York YO10 5DD, United Kingdom.
authors.forename: Vanda
authors.initials: V
authors.lastname: Gunning
authors.affiliation: Centre for Novel Agricultural Products (V.G., K.T., H.S., E.J.J., E.L.R., N.C.B.) and Department of Biology (A.S.B., J.R.P.), University of York, York YO10 5DD, United Kingdom.
authors.forename: Kyriakos
authors.initials: K
authors.lastname: Tzafestas
authors.affiliation: Centre for Novel Agricultural Products (V.G., K.T., H.S., E.J.J., E.L.R., N.C.B.) and Department of Biology (A.S.B., J.R.P.), University of York, York YO10 5DD, United Kingdom.
authors.forename: Helen
authors.initials: H
authors.lastname: Sparrow
authors.affiliation: Centre for Novel Agricultural Products (V.G., K.T., H.S., E.J.J., E.L.R., N.C.B.) and Department of Biology (A.S.B., J.R.P.), University of York, York YO10 5DD, United Kingdom.
authors.forename: Emily J
authors.initials: EJ
authors.lastname: Johnston
authors.affiliation: Centre for Novel Agricultural Products (V.G., K.T., H.S., E.J.J., E.L.R., N.C.B.) and Department of Biology (A.S.B., J.R.P.), University of York, York YO10 5DD, United Kingdom.
authors.forename: Andrew S
authors.initials: AS
authors.lastname: Brentnall
authors.affiliation: Centre for Novel Agricultural Products (V.G., K.T., H.S., E.J.J., E.L.R., N.C.B.) and Department of Biology (A.S.B., J.R.P.), University of York, York YO10 5DD, United Kingdom.
authors.forename: Jennifer R
authors.initials: JR
authors.lastname: Potts
authors.affiliation: Centre for Novel Agricultural Products (V.G., K.T., H.S., E.J.J., E.L.R., N.C.B.) and Department of Biology (A.S.B., J.R.P.), University of York, York YO10 5DD, United Kingdom liz.rylott@york.ac.uk neil.bruce@york.ac.uk.
authors.forename: Elizabeth L
authors.initials: EL
authors.lastname: Rylott
authors.affiliation: Centre for Novel Agricultural Products (V.G., K.T., H.S., E.J.J., E.L.R., N.C.B.) and Department of Biology (A.S.B., J.R.P.), University of York, York YO10 5DD, United Kingdom liz.rylott@york.ac.uk neil.bruce@york.ac.uk.
authors.forename: Neil C
authors.initials: NC
authors.lastname: Bruce
references.citation: Plant Physiol. 1993 Jul;102(3):803-10
references.pmid: 8278534
references.citation: Chemosphere. 2001 Aug;44(5):1259-64
references.pmid: 11513416
references.citation: Plant Physiol. 1999 Dec;121(4):1291-8
references.pmid: 10594116
references.citation: Chemosphere. 2007 Jun;68(6):1050-7
references.pmid: 17368510
references.citation: Plant Physiol. 2005 Jun;138(2):858-69
references.pmid: 15923336
references.citation: Plant J. 2008 Dec;56(6):963-74
references.pmid: 18702669
references.citation: Chem Res Toxicol. 2001 Jan;14(1):62-70
references.pmid: 11170509
references.citation: Plant Cell Environ. 2001 May;24(5):557-63
references.pmid: 11706851
references.citation: Int J Phytoremediation. 2007 Sep-Oct;9(5):385-401
references.pmid: 18246725
references.citation: Nature. 2003 Sep 25;425(6956):393-7
references.pmid: 14508488
references.citation: Org Biomol Chem. 2006 Apr 7;4(7):1252-60
references.pmid: 16557313
references.citation: Anal Biochem. 2007 Apr 1;363(1):58-69
references.pmid: 17288982
references.citation: Nat Biotechnol. 2001 Dec;19(12):1168-72
references.pmid: 11731787
references.citation: Curr Opin Plant Biol. 1998 Jun;1(3):258-66
references.pmid: 10066594
references.citation: Environ Pollut. 2010 Feb;158(2):470-5
references.pmid: 19782446
references.citation: Arabidopsis Book. 2010;8:e0131
references.pmid: 22303257
references.citation: Int J Phytoremediation. 2007 Jan-Feb;9(1):15-30
references.pmid: 18246712
references.citation: Biotechnol Genet Eng Rev. 2001;18:171-217
references.pmid: 11530688
references.citation: Methods Enzymol. 2005;401:169-86
references.pmid: 16399386
references.citation: Plant Mol Biol. 1992 Dec;20(6):1203-7
references.pmid: 1463857
references.citation: J Exp Bot. 2009;60(4):1207-18
references.pmid: 19174456
references.citation: Rev Environ Contam Toxicol. 1999;161:1-156
references.pmid: 10218448
references.citation: Drug Metab Rev. 2011 May;43(2):266-80
references.pmid: 21425939
references.citation: New Phytol. 2011 Oct;192(2):405-13
references.pmid: 21729248
references.citation: Environ Sci Technol. 2008 Oct 1;42(19):7405-10
references.pmid: 18939578
references.citation: Trends Biotechnol. 2009 Feb;27(2):73-81
references.pmid: 19110329
references.citation: Chemosphere. 2008 Oct;73(5):657-62
references.pmid: 18774158
references.citation: Environ Sci Pollut Res Int. 1998;5(4):202-8
references.pmid: 19002633
references.citation: J Biol Chem. 2009 Aug 7;284(32):21249-56
references.pmid: 19520850
references.citation: Proc Natl Acad Sci U S A. 2007 Oct 23;104(43):16822-7
references.pmid: 17940033
references.citation: Appl Environ Microbiol. 1998 Aug;64(8):2864-8
references.pmid: 9687442
references.citation: Curr Opin Biotechnol. 2011 Jun;22(3):434-40
references.pmid: 21094036
references.citation: Methods. 2001 Dec;25(4):402-8
references.pmid: 11846609
references.citation: Mol Genet Genomics. 2008 Apr;279(4):339-57
references.pmid: 18270741
references.citation: FEBS Lett. 2006 Nov 27;580(27):6384-90
references.pmid: 17097087
references.citation: Plant Physiol. 2009 Sep;151(1):253-61
references.pmid: 19605548
references.citation: Environ Sci Pollut Res Int. 2004;11(5):331-9
references.pmid: 15506637
references.citation: Plant Physiol. 2003 Nov;133(3):1397-406
references.pmid: 14551330
references.citation: Chemosphere. 2006 May;63(8):1280-90
references.pmid: 16352328
references.citation: J Exp Bot. 2008;59(14):3857-68
references.pmid: 18849295
references.citation: Drug Metab Dispos. 2009 Jun;37(6):1242-50
references.pmid: 19282395
references.citation: Nat Biotechnol. 2006 Feb;24(2):216-9
references.pmid: 16429147
references.citation: Protein Expr Purif. 2005 May;41(1):207-34
references.pmid: 15915565
references.citation: Plant J. 2007 Mar;49(5):878-88
references.pmid: 17316176