Sharma R, Gupta S, Singhal S S, Ahmad H, Haque A, Awasthi Y C
Department of Human Biological Chemistry and Genetics, University of Texas Medical Branch, Galveston 77550.
Biochem J. 1991 Apr 15;275 ( Pt 2)(Pt 2):507-13. doi: 10.1042/bj2750507.
Glutathione S-transferase (GST) isoenzymes of human pancreas were purified, characterized and evaluated for their possible role in the metabolism of ethanol. Human pancreas has at least two GST isoenzymes belonging to the Alpha class (pI 8.8 and 8.1), one belonging to the Mu class (pI 6.4) and one belonging to the Pi class (pI 4.9). During the purification of GSTs from pancreas as well as from heart, liver, lung, brain and muscle, the fatty acid ethyl ester synthase (FAEES) activity was monitored in order to evaluate the role of GSTs in metabolism of ethanol, as suggested in earlier studies. Both t.l.c. and h.p.l.c. were used to identify ethyl oleate in reaction mixtures to monitor FAEES activity. During the purification of GSTs with the use of affinity chromatography on GSH linked to epoxy-activated Sepharose 6B, FAEES and GST activities from each of these tissues segregated independently. Purified GST isoenzymes from these tissues did not exhibit any FAEES activity. Antibodies raised against Pi-class GST, as expected, immunoprecipitated most of the GST activity of brain and heart without precipitating FAEES activity. These results suggest that human GST isoenzymes belonging to the Alpha, Mu and Pi classes do not express FAEES activity. The independent segregation of GST and FAEES activities was further demonstrated by monitoring GST activity during the purification of FAEES from pancreas. It was found that purified FAEES had no GST activity towards 1-chloro-2,4-dinitrobenzene and a number of other electrophilic substrates. Results of these studies demonstrate that FAEES and GSTs are distinct proteins.
对人胰腺的谷胱甘肽S-转移酶（GST）同工酶进行了纯化、表征，并评估了它们在乙醇代谢中可能发挥的作用。人胰腺至少有两种属于α类的GST同工酶（pI 8.8和8.1），一种属于μ类（pI 6.4），一种属于π类（pI 4.9）。在从胰腺以及心脏、肝脏、肺、脑和肌肉中纯化GST的过程中，按照早期研究的建议，监测了脂肪酸乙酯合酶（FAEES）的活性，以评估GST在乙醇代谢中的作用。采用薄层层析（t.l.c.）和高效液相色谱（h.p.l.c.）来鉴定反应混合物中的油酸乙酯，以监测FAEES活性。在用与环氧活化的琼脂糖6B偶联的谷胱甘肽进行亲和层析纯化GST的过程中，这些组织中的每一个组织的FAEES和GST活性都是独立分离的。从这些组织中纯化得到的GST同工酶没有表现出任何FAEES活性。如预期的那样，针对π类GST产生的抗体免疫沉淀了大脑和心脏的大部分GST活性，而没有沉淀FAEES活性。这些结果表明，属于α、μ和π类的人GST同工酶不表达FAEES活性。在从胰腺纯化FAEES的过程中监测GST活性，进一步证明了GST和FAEES活性的独立分离。研究发现，纯化后的FAEES对1-氯-2,4-二硝基苯和许多其他亲电底物没有GST活性。这些研究结果表明，FAEES和GST是不同的蛋白质。
