Glutathione S-transferases of the yeast Yarrowia lipolytica have unusually large molecular mass.
Two similar glutathione S-transferases (GSTs), which do not bind to glutathione- or S-hexylglutathione-agarose affinity resins, have been purified from the yeast Yarrowia lipolytica. An approx. 400-fold purification was obtained by a combination of DEAE-Sephadex, phenyl-Sepharose, hydroxyapatite and Mono-Q anion-exchange chromatography. The native molecular mass of both proteins was estimated as approx. 110 kDa by both Superose-12 gel-filtration chromatography and non-denaturing electrophoresis. SDS/PAGE indicated a subunit mass of 50 kDa. Reverse-phase HPLC of purified proteins gave a single, well-resolved, peak, suggesting that the proteins are homodimers. Identical behaviour on HPLC, native electrophoresis and SDS/PAGE, N-terminal sequencing, sensitivity to a panel of inhibitors and identical specific activities with 1-chloro-2,4-dinitrobenzene as substrate suggest that the two isoenzymes are very similar. The enzymes do not immunoblot with antisera to any of the main GST classes, and N-terminal sequencing suggests no clear relationship with previously characterized enzymes, such as that of the fungus, Phanerochaete chrysosporium [Dowd, Buckley and Sheehan (1997) Biochem. J. 324, 243-248]. It is possible that the two isoenzymes arise as a result of post-translational modification of a single GST isoenzyme.
Department of Biochemistry, University College Cork, Lee Maltings, Prospect Row, Cork, Republic of Ireland.
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Foley
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Sheehan
D000595:Amino Acid Sequence; D002850:Chromatography, Gel; D005656:Fungal Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D005982:Glutathione Transferase / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011487:Protein Conformation; D004718:Saccharomycetales / Q000201:enzymology*; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D005656:Fungal Proteins; D005982:Glutathione Transferase