Nakano Hiroaki, Ogura Kenichiro, Takahashi Eriko, Harada Tomokazu, Nishiyama Takahito, Muro Kei, Hiratsuka Akira, Kadota Shigetoshi, Watabe Tadashi
Corporate, Scientific and Regulatory Affairs Division, Tobacco Headquarters, Japan Tobacco Inc., Tokyo.
Drug Metab Pharmacokinet. 2004 Jun;19(3):216-26. doi: 10.2133/dmpk.19.216.
Regioselective sulfation of the phytoestrogens daidzein (DZ, 7,4'-dihydroxyisoflavone) and genistein (GS, 5,7,4'-trihydroxyisoflavone) was investigated using human liver cytosol and purified recombinant human sulfotransferase (SULT) isoforms, SULT1A1, SULT1A3, SULT2A1, and SULT1E1. 7-Position-preferential sulfation of DZ and GS was observed in human hepatic cytosols from 3 male and 3 female subjects. Average ratios for 7- to 4'-sulfate formation were 4.5:1 from DZ and 8.4:1 from GS in these human liver cytosols. Apparent K(m) values for the 7- and 4'-sulfation of DZ and GS by these cytosols were similar and in a range from 0.46 to 0.66 microM. All recombinant human SULTs had activity for 7- and 4'-sulfation of these phytoestrogens except for 7-sulfating activity of SULT1A3. SULT1A1 and SULT1E1 exhibited much higher catalytic efficiency, k(cat)/K(m), for 7- and 4'-sulfation of these substrates than did the other two, SULT1A3 and SULT2A1. SULT1A1 showed K(m) values of 0.47 and 0.52 microM for the mono-sulfation of DZ and GS, respectively, which were very similar to those of human cytosol. The observed k(cat)/K(m) indicated that SULT1A1 catalyzed 7-sulfation of DZ and GS at rates 4.4- and 8.8-fold higher, respectively, than such 4'-sulfation. However, with SULT1E1, catalytic efficiency was very similar for the sulfation of both positions. These data strongly suggest that SULT1A1 plays a major role in monosulfation of the phytoestrogens and determines the regioselectivity of sulfation in human hepatic cytosol. A kinetic study for 7,4'-disulfate formation of DZ and GS from their 7- and 4'-monosulfates indicated that SULT1E1 most efficiently catalyzed both reactions among human SULTs.
利用人肝细胞溶胶和纯化的重组人磺基转移酶（SULT）同工型SULT1A1、SULT1A3、SULT2A1和SULT1E1，研究了植物雌激素大豆苷元（DZ，7,4'-二羟基异黄酮）和染料木黄酮（GS，5,7,4'-三羟基异黄酮）的区域选择性硫酸化。在3名男性和3名女性受试者的人肝细胞溶胶中观察到DZ和GS的7位优先硫酸化。在这些人肝细胞溶胶中，DZ生成7-硫酸酯与4'-硫酸酯的平均比率为4.5:1，GS为8.4:1。这些细胞溶胶对DZ和GS的7位和4'-位硫酸化的表观K(m)值相似，范围为0.46至0.66微摩尔。除SULT1A3的7位硫酸化活性外，所有重组人SULT对这些植物雌激素的7位和4'-位硫酸化均有活性。与另外两种SULT1A3和SULT2A1相比，SULT1A1和SULT1E1对这些底物的7位和4'-位硫酸化表现出更高的催化效率k(cat)/K(m)。SULT1A1对DZ和GS单硫酸化的K(m)值分别为0.47和0.52微摩尔，与人细胞溶胶的值非常相似。观察到的k(cat)/K(m)表明，SULT1A1催化DZ和GS的7位硫酸化的速率分别比4'-位硫酸化高4.4倍和8.8倍。然而，对于SULT1E1，两个位置硫酸化的催化效率非常相似。这些数据强烈表明，SULT1A1在植物雌激素的单硫酸化中起主要作用，并决定了人肝细胞溶胶中硫酸化的区域选择性。一项关于从DZ和GS的7-和4'-单硫酸酯形成7,4'-二硫酸酯的动力学研究表明，在人SULT中，SULT1E1最有效地催化这两个反应。
