Shimada T, Yamazaki H, Oda Y, Hiratsuka A, Watabe T, Guengerich F P
Osaka Prefectural Institute of Public Health, Japan.
Chem Res Toxicol. 1996 Jan-Feb;9(1):333-40. doi: 10.1021/tx950125v.
A newly developed tester Salmonella typhimurium NM5004 strain was constructed by introducing a plasmid containing both rat GSH S-transferase (GST) 5-5 cDNA and the umuC"lacZ operon into the host strain Salmonella typhimurium TA1535 and used to examine whether or not GST modified the genotoxic activities of several dihaloalkanes and other compounds. Twenty-nine chemicals that were suggested to be conjugated by GST were compared with regard to their abilities to induce umu gene expression and cause cytotoxicity responses in both the NM5004 strain and the original tester strain (S. typhimurium TA1535/pSK1002, which is devoid of GST activity toward 1,2-epoxy-3-(4'-nitrophenoxy)propane). Ten chemicals--1,2-dibromoethane,N-(2,3-epoxypropyl)phthalimide, 1,3-dichloroacetone, CH2I2, 1,2-epoxy-3-phenoxypropane, 2,3-epoxypropyl p-methoxyphenyl ether, 1-bromo-2-chloroethane, 1-bromo-2,3-dichloropropane, CH2BrCl, and CH2Br2--were found to enhance induction of umu gene expression in the NM5004 strain as compared with the TA1535/pSK1002 strain. 1,2-Epoxy-3-(4'-nitrophenoxy)propane and 2,3-dibromo-1-chloropropane were inactivated by GST 5-5 in the NM5004 tester strain, although these chemicals were cytotoxic in both tester strains. Roles of GST 5-5 were also examined for the inactivation of reactive metabolites of several procarcinogens that were formed through oxidation by liver microsomes of polychlorinated biphenyl-treated rats. The results suggest that reactive metabolites (possibly epoxides) of aflatoxin B1, sterigmatocystin, 1,2-dihydro-1,2-dihydroxy-6-aminochrysene, and (+)- and (-)-enantiomers of 7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene could be trapped as inactivated GSH conjugates in the NM5004 strain. High-performance liquid chromatographic analysis suggested that exo-aflatoxin B1-8,9-oxide--GSH conjugate was formed during the oxidation of aflatoxin B1 by rat and human liver microsomes in the presence of GSH and several GST enzymes including purified rat theta class GST Yrs-Yrs and rat liver GST (a mixture of alpha and mu class enzymes). Thus, the present results support the view that the theta class rat GST 5-5 enzyme participates in the activation and inactivation of potential environmental carcinogenic chemicals. This newly developed NM5004 tester strain is of use in the elucidation of roles of GST 5-5 in transformations.
通过将含有大鼠谷胱甘肽S-转移酶（GST）5-5 cDNA和umuC"lacZ操纵子的质粒导入宿主菌株鼠伤寒沙门氏菌TA1535，构建了一种新开发的测试菌株鼠伤寒沙门氏菌NM5004，并用于检测GST是否改变了几种二卤代烷和其他化合物的遗传毒性活性。将29种被认为可被GST共轭的化学物质在诱导umu基因表达的能力以及在NM5004菌株和原始测试菌株（鼠伤寒沙门氏菌TA1535/pSK1002，其对1,2-环氧-3-(4'-硝基苯氧基)丙烷没有GST活性）中引起细胞毒性反应方面进行了比较。与TA1535/pSK1002菌株相比，发现10种化学物质——1,2-二溴乙烷、N-(2,3-环氧丙基)邻苯二甲酰亚胺、1,3-二氯丙酮、CH2I2、1,2-环氧-3-苯氧基丙烷、2,3-环氧丙基对甲氧基苯基醚、1-溴-2-氯乙烷、1-溴-2,3-二氯丙烷、CH2BrCl和CH2Br2——能增强NM5004菌株中umu基因表达的诱导。1,2-环氧-3-(4'-硝基苯氧基)丙烷和2,3-二溴-1-氯丙烷在NM5004测试菌株中被GST 5-5灭活，尽管这些化学物质在两种测试菌株中均具有细胞毒性。还研究了GST 5-5在灭活几种经多氯联苯处理的大鼠肝微粒体氧化形成的前致癌物的活性代谢产物中的作用。结果表明，黄曲霉毒素B1、杂色曲霉素、1,2-二氢-1,2-二羟基-6-氨基芘以及7,8-二羟基-7,8-二氢苯并[a]芘的(+)和(-)对映体的活性代谢产物（可能是环氧化物）可在NM5004菌株中作为灭活的谷胱甘肽共轭物被捕获。高效液相色谱分析表明，在谷胱甘肽和几种GST酶（包括纯化的大鼠θ类GST Yrs-Yrs和大鼠肝GST（α和μ类酶的混合物））存在的情况下，大鼠和人肝微粒体氧化黄曲霉毒素B1期间形成了外消旋黄曲霉毒素B1-8,9-氧化物-GSH共轭物。因此，目前的结果支持这样一种观点，即大鼠θ类GST 5-5酶参与了潜在环境致癌化学物质的激活和灭活。这种新开发的NM5004测试菌株可用于阐明GST 5-5在转化中的作用。
