Identification of multiple glutathione S-transferases from Daphnia magna.

1. Six anionic glutathione S-transferases (GST) were purified from the crustacean, Daphnia magna, by means of affinity chromatography, that are responsible for ca. 40% of cytosolic GST activity. 2. Electrophoresis in the presence of sodium dodecyl sulfate (SDS) revealed the presence of three proteins, with molecular weights of 27,500, 28,000, and 30,200. 3. Separation under nondenaturing conditions revealed six proteins, all of which exhibited GST activity, with molecular weights ranging from 55,000 to 61,700. 4. Ethacrynic acid is a competitive inhibitor of activity towards CDNB of all six GSTs, binding each with similar affinities. 5. Chlorinated phenols are also competitive inhibitors of the enzyme, with the degree of inhibition being directly correlated with the lipophilicity of the compounds. 6. Analysis of inhibition of separated isoforms reveals that form 4 is most strongly inhibited by these chlorinated phenols, with forms 5 and 6 being inhibited to a lesser degree.