Glucuronidation of acetaminophen catalyzed by multiple rat phenol UDP-glucuronosyltransferases
Glucuronidation of acetaminophen catalyzed by multiple rat phenol UDP-glucuronosyltransferases
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DOI：
10.1124/dmd.30.3.324
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发表时间：
2002-03-01
影响因子：
3.9
通讯作者：
Ritter, JK
中科院分区：
文献类型：
--
作者：
Kessler, FK;Kessler, MR;Ritter, JK
大鼠葡萄糖醛酸对乙酰氨基酚（APAP）发生率降低，对APAP诱导的肝毒性易感性增加。这种缺陷被认为与udp -葡萄糖醛基转移酶（UGT） 1A6有关，该酶在Gunn大鼠中不起作用，但目前尚不清楚其他1A家族成员是否也参与其中。在人类中，已知有两种1A亚型是活跃的（1A6和1A9），但1A6形式的表观K-m （2mm）低25倍。大鼠肝微粒体APAP UGT活性可由体内β -萘黄酮或奥替普拉诱导，其作用与1A6和1A7的诱导有关。为了确定1A7相对于其他1A形式在APAP糖醛酸化中的可能作用，编码UGTs 1A1、1A5、1A6、1A7和1A8的cdna在人胚胎肾细胞中表达，并通过定量免疫印迹测定制备的膜组分中表达酶的含量。在2.5 mM APAP下，1A7蛋白的比活性最高（2.8 nmol/min/nmol），其次是1A6 （1.1 nmol/min/nmol）和1A8 （0.27 nmol/min/nmol）。1A1和1A5基本上没有活性。动力学比较表明，1A7具有与1A6相似的表观K（分别为4.7 mM和3.9 mM），但催化活性高2.4倍。这些数据表明，在大鼠中，1A7在APAP糖醛酸化中起主要作用，并有助于防止APAP诱导的肝毒性。Gunn大鼠肝细胞中存在显著残余的apap -葡萄糖醛酸活性，进一步强调了除1A6外其他UGTs的参与，表明UGT2家族成员的活性未知。
Gunn rats glucuronidate acetaminophen (APAP) at reduced rates and show increased susceptibility to APAP-induced hepatotoxicity. This defect is presumed to involve UDP-glucuronosyltransferase (UGT) 1A6, which is nonfunctional in Gunn rats, but it is currently unclear whether other 1A family members are also involved. In humans, two 1A isoforms are known to be active (1A6 and 1A9) but 1A6 form has a 25-fold lower apparent K-m (2 mM). Rat liver microsomal APAP UGT activity is induced by in vivo treatment with beta-naphthoflavone or oltipraz, an effect correlating with induction of 1A6 and 1A7. To address a possible role of 1A7 in APAP glucuronidation relative to other 1A forms, cDNAs encoding UGTs 1A1, 1A5, 1A6, 1A7, and 1A8 were expressed in human embryonic kidney cells and the contents of expressed enzyme in prepared membrane fractions determined by quantitative immunoblotting. At 2.5 mM APAP, 1A7 showed the highest specific activity (2.8 nmol/min/nmol 1A7 protein), followed by 1A6 (1.1 nmol/min/nmol), and 1A8 (0.27 nmol/min/nmol). 1A1 and 1A5 were essentially inactive. Kinetic comparisons indicated 1A7 had a similar apparent K, as 1A6 (4.7 versus 3.9 mM, respectively) but a 2.4-fold higher catalytic activity. These data suggest that in rats, 1A7 plays a major role in APAP glucuronidation and contributes to protection against APAP-induced hepatotoxicity. The involvement of other UGTs besides 1A6 is further underscored by the presence of significant residual APAP-glucuronidating activity by Gunn rat hepatocytes, indicating the activity of an unknown UGT2 family member.