Purification and properties of anionic glutathione S-transferase from bovine ciliary body.
An anionic glutathione S-transferase was purified from bovine ciliary body by DEAE-agarose chromatography and affinity chromatography on GSH-agarose and Orange A. The enzyme accounts for about 25% of total soluble glutathione S-transferase activity of the tissue. The purified enzyme has a molecular mass of about 50,000 Da and is composed of two identical subunits of about 25,000 Da. The enzyme has a pI of 5.8. The enzyme conjugates GSH with 1-chloro-2,4-dinitrobenzene, p-nitrobenzyl chloride, 1,2-epoxy-3-(p-nitrophenyl)propane, 1,2-dinitrobenzene and 3,4-dinitrobenzoic acid. The Km values for 1-chloro-2,4-dinitrobenzene and GSH are 0.40 mM and 0.57 mM respectively. Haematin is a non-competitive inhibitor (Ki = 4.5 microM) when tested with various concentrations of 1-chloro-2,4-dinitrobenzene. The enzyme shows no glutathione peroxidase activity with either H2O2 or cumene peroxide as substrate. On the basis of substrate specificities, pI values, amino acid composition and peptide maps, it is concluded that the ciliary-body enzyme is probably identical with the anionic form of glutathione S-transferase from bovine lens and liver.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001663:Bilirubin / Q000494:pharmacology; D002417:Cattle; D002924:Ciliary Body / Q000201:enzymology*; D005982:Glutathione Transferase / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D006427:Hemin / Q000494:pharmacology; D007525:Isoelectric Focusing; D008970:Molecular Weight; D010449:Peptide Mapping; D011524:Protoporphyrins / Q000494:pharmacology
D000596:Amino Acids; D011524:Protoporphyrins; D006427:Hemin; C028025:protoporphyrin IX; D005982:Glutathione Transferase; D001663:Bilirubin