Hu X, Srivastava S K, Xia H, Awasthi Y C, Singh S V
Cancer Research Laboratory, Mercy Cancer Institute, Mercy Hospital of Pittsburgh, Pittsburgh, Pennsylvania 15219, USA.
J Biol Chem. 1996 Dec 20;271(51):32684-8. doi: 10.1074/jbc.271.51.32684.
The kinetics of the conjugation of glutathione (GSH) with anti-7beta, 8alpha-dihydroxy-9alpha,10alpha-oxy-7,8,9, 10-tetrahydrobenzo(a)pyrene (anti-BPDE) catalyzed by GSH S-transferase (GST) isoenzymes purified from the liver and forestomach of female A/J mouse has been investigated. The GST isoenzymes studied included an alpha class isoenzyme of forestomach (GST 9.5), alpha class hepatic isoenzymes mGSTA3-3 and mGSTA4-4, pi class hepatic isoenzyme mGSTP1-1, and mu class hepatic isoenzyme mGSTM1-1. When the concentration of (+)-anti-BPDE was varied (5-120 microM) at a fixed GSH concentration (2 mM), linear Lineweaver-Burk plots were observed for each isoenzyme. The kcat values for GST 9.5, mGSTA3-3, mGSTP1-1, mGSTM1-1, and mGSTA4-4 were 2.0, 0.02, 0.40, 0. 05, and 0.01 s-1, respectively, with corresponding Km values of 16, 12, 29, 27, and 49 microM. The catalytic efficiency (kcat/Km) of GST 9.5 in the conjugation of GSH with (+)-anti-BPDE, which is believed to be the ultimate carcinogenic metabolite of benzo(a)pyrene, was about 9-625-fold higher as compared with other mouse GST isoenzymes. These results indicate that GST 9.5 of forestomach is different among mammalian alpha class GSTs because (+)-anti-BPDE has been shown to be a poor substrate for alpha class rat or human GST isoenzymes. The catalytic efficiency of GST 9.5 was approximately 4.5-fold higher than that of pi class human isoenzyme (hGSTP1-1), which among human GSTs is reported to be most efficient in the detoxification of (+)-anti-BPDE. Unlike rat GST isoenzymes, linear Lineweaver-Burk plots were observed for mouse GSTs when GSH was used as a variable substrate. The catalytic efficiencies of the mouse GSTs toward (+)-anti-BPDE were about 2-20-fold higher as compared with the (-)-enantiomer of anti-BPDE. The results of the present study suggest that GST 9.5 may play an important role in the detoxification of (+)-anti-BPDE.
对从雌性A/J小鼠肝脏和前胃中纯化得到的谷胱甘肽S-转移酶（GST）同工酶催化谷胱甘肽（GSH）与反式-7β, 8α-二羟基-9α,10α-环氧-7,8,9,10-四氢苯并(a)芘（反式-BPDE）结合反应的动力学进行了研究。所研究的GST同工酶包括前胃的α类同工酶（GST 9.5）、肝脏α类同工酶mGSTA3-3和mGSTA4-4、肝脏π类同工酶mGSTP1-1以及肝脏μ类同工酶mGSTM1-1。当在固定的GSH浓度（2 mM）下改变（+）-反式-BPDE的浓度（5 - 120 μM）时，观察到每种同工酶的Lineweaver-Burk图呈线性。GST 9.5、mGSTA3-3、mGSTP1-1、mGSTM1-1和mGSTA4-4的kcat值分别为2.0、0.02、0.40、0.05和0.01 s-1，相应的Km值分别为16、12、29、27和49 μM。GST 9.5催化GSH与（+）-反式-BPDE结合反应的催化效率（kcat/Km），据信（+）-反式-BPDE是苯并(a)芘的最终致癌代谢物，与其他小鼠GST同工酶相比高约9 - 625倍。这些结果表明，前胃的GST 9.5在哺乳动物α类GST中有所不同，因为已表明（+）-反式-BPDE是大鼠或人类α类GST同工酶的不良底物。GST 9.5的催化效率比人类π类同工酶（hGSTP1-1）高约4.5倍，据报道在人类GST中hGSTP1-1对（+）-反式-BPDE的解毒作用最有效。与大鼠GST同工酶不同，当将GSH用作可变底物时，观察到小鼠GST的Lineweaver-Burk图呈线性。小鼠GST对（+）-反式-BPDE的催化效率比反式-BPDE的（-）-对映体高约2 - 20倍。本研究结果表明，GST 9.5可能在（+）-反式-BPDE的解毒中起重要作用。
