Glutathione S-transferases from the white-rot fungus, Phanerochaete chrysosporium.
A glutathione S-transferase (GST) was purified to homogeneity from the white-rot fungus, Phanerochaete chrysosporium, by affinity chromatography on glutathione-agarose followed by Mono-Q ion-exchange FPLC. This protein immunoblotted with antisera to rat Theta class GST 5-5 and also showed N-terminal sequence similarity to the Theta class, including the presence of a conserved serine residue that has been specifically implicated in catalysis in this class [Wilce, Board, Feil and Parker (1995) EMBO J. 14, 2133-2143] and other residues conserved in plant sequences. Catalytic activity was found to be highly labile in the purified protein, although preliminary evidence for activity (approx. 120 m-units/mg) with 1,2-epoxy-3-(p-nitrophenoxy)propane was obtained in some preparations. The enzyme seems to be a dimer with a subunit molecular mass of 25 kDa by SDS/PAGE. The native molecular masses estimated by non-denaturing electrophoresis and by Superose-12 gel filtration were 58 and 45 kDa respectively. A second protein purified in this study also gave low level of activity with 1,2-epoxy-3-(p-nitrophenoxy)propane and had a subunit molecular mass of 28 kDa (native size 62-63 kDa), but did not immunoblot with any GST class and seemed to be N-terminally blocked.
Dowd
C A
CA
Department of Biochemistry, University College Cork, Lee Maltings, Prospect Row, Cork, Ireland.
Buckley
C M
CM
Sheehan
D
D
D000363:Agaricales / Q000201:enzymology*; D000595:Amino Acid Sequence; D000818:Animals; D001419:Bacteria / Q000201:enzymology; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D017124:Conserved Sequence; D004591:Electrophoresis, Polyacrylamide Gel; D005399:Fishes; D005658:Fungi / Q000201:enzymology; D005982:Glutathione Transferase / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D007313:Insecta; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D051381:Rats; D017386:Sequence Homology, Amino Acid; D012694:Serine
D046911:Macromolecular Substances; D012694:Serine; D005982:Glutathione Transferase