Purification of a new acidic glutathione S‐transferase, GST‐Yn1 Yn1, with a high leukotriene‐C4 synthase activity from rat brain

Abstract

A new acidic form of glutathione S‐transferase (GST, pI 6.2) was purified from rat brain by S‐hexylglutathione affinity chromatography followed by chromatofocusing. This form occupied 20–25% of the total activity bound to the affinity column. It had a molecular mass (subunit 26 kDa) similar to that of a major GST form of rat testis (MT or 6‐6) on sodium dodecyl sulfate/polyacrylamide gel electrophoresis. However, it differed from the MT in isoelectric point, activity towards 1,2‐dichloro‐4‐nitrobenzene and immunological properties. On two‐dimensional gel electrophoresis the brain form gave a spot which was identical in molecular mass, isoelectric point and immunological properties to a less acidic one (Yn1) of two spots (Yn1 and Yn2) of the testis GST‐MT. Therefore, the brain acidic form is a homodimer, and named GST‐Yn1 Yn1. The activity was inhibited by sulfasalazine, an inhibitor of leukotriene‐C4 synthase. This form (GST‐Yn1Yn1) showed the highest leukotriene‐C4 synthase activity, 496 nmol/mg protein in 5 min, among nine cytosolic GST isoenzymes from the rat. The Km values for leukotriene A4 and glutathione were 26 μM and 3.5 mM respectively. A major GST form of rat brain, occupying about 40% of the total activity, was identical with GST‐P (7–7) purified from rat liver bearing preneoplastic hyperplastic nodules and localized at astroglias. GST‐P also showed the significant leukotriene‐C4 synthase activity, 67.2 nmol/mg protein in 5 min, but the Km for leukotriene A4 was 100 μM, fourfold higher than that of GST‐Yn1 Yn1. These results suggest that mainly GST‐Yn1 Yn1 may be involved in leukotriene‐C4 synthesis in rat brain. Copyright © 1987, Wiley Blackwell. All rights reserved