Sieg A, van Hees G P, Heirwegh K P
J Clin Invest. 1982 Feb;69(2):347-57. doi: 10.1172/jci110458.
glucuronyl transfer (a) from UDP-glucuronic acid to bilirubin monoglucuronide, catalyzed by a microsomal UDP-glucuronyltransferase, and (b) from one molecule of bilirubin monoglucuronide to another (transglucuronidation), catalyzed by an enzyme present in liver plasma membranes. The evidence regarding the role of the latter enzyme for in vivo formation of bilirubin diglucuronide is conflicting. We therefore decided to reexamine the transglucuronidation reaction in plasma membranes and to study the conversion of bilirubin monoglucuronide to diglucuronide in vivo. Purified bilirubin monoglucuronide was incubated with homogenates and plasma membrane-enriched fractions from liver of Wistar and Gunn rats. Stoichiometric formation of bilirubin and bilirubin diglucuronide out of 2 mol of bilirubin monoglucuronide was paralleled by an increase of the IIIalpha- and XIIIalpha-isomers of the bilirubin aglycone, thus showing that dipyrrole exchange, not transglucuronidation, is the underlying mechanism. Complete inhibition by ascorbic acid probably reflects intermediate formation of free radicals of dipyrrolic moieties. The reaction was nonenzymic because it proceeded independently of the protein concentration and heat denaturation of the plasma membranes did not result in decreased conversion rates. Collectively, these findings show spontaneous, nonenzymic dipyrrole exchange when bilirubin monoglucuronide is incubated in the presence of rat liver plasma membranes. Because bilirubin glucuronides present in biological fluids contain exclusively the bilirubin-IXalpha aglycone, formation of the diglucuronide from the monoglucuronide by dipyrrole exchange does not occur in vivo. Rapid excretion of unchanged bilirubin monoglucuronide in Gunn rat bile after injection of the pigment provides confirmatory evidence for the absence of a UDP-glucuronic acid-independent process.
关于胆红素单葡萄糖醛酸酯转化为双葡萄糖醛酸酯，已提出两条途径：(a) 由微粒体UDP-葡萄糖醛酸基转移酶催化，将UDP-葡萄糖醛酸的葡萄糖醛酸基转移至胆红素单葡萄糖醛酸酯；(b) 由肝细胞膜中存在的一种酶催化，从一分子胆红素单葡萄糖醛酸酯转移至另一分子（转葡萄糖醛酸化）。关于后一种酶在体内胆红素双葡萄糖醛酸酯形成中所起作用的证据相互矛盾。因此，我们决定重新研究细胞膜中的转葡萄糖醛酸化反应，并研究胆红素单葡萄糖醛酸酯在体内向双葡萄糖醛酸酯的转化。将纯化的胆红素单葡萄糖醛酸酯与Wistar大鼠和Gunn大鼠肝脏的匀浆及富含细胞膜的组分一起孵育。两分子胆红素单葡萄糖醛酸酯按化学计量生成胆红素和胆红素双葡萄糖醛酸酯，同时胆红素苷元的IIIα-和XIIIα-异构体增加，这表明潜在机制是二吡咯交换而非转葡萄糖醛酸化。抗坏血酸的完全抑制作用可能反映了二吡咯部分自由基的中间形成。该反应是非酶促反应，因为其进行与蛋白质浓度无关，且细胞膜的热变性不会导致转化率降低。总体而言，这些发现表明，当胆红素单葡萄糖醛酸酯在大鼠肝细胞膜存在的情况下孵育时，会发生自发的非酶促二吡咯交换。由于生物流体中存在的胆红素葡萄糖醛酸酯仅含有胆红素-IXα苷元，因此在体内不会通过二吡咯交换由单葡萄糖醛酸酯形成双葡萄糖醛酸酯。注射色素后，Gunn大鼠胆汁中未变化的胆红素单葡萄糖醛酸酯迅速排泄，这为不存在不依赖UDP-葡萄糖醛酸的过程提供了确证。
