Biliverdin-IXa Reductase and Biliverdin-IXP Reductase from Human Liver
Biliverdin-IXa Reductase and Biliverdin-IXP Reductase from Human Liver
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发表时间：
1994
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通讯作者：
T. Yamaguchif;Yasuo KomodaP;H. Nakajima
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作者：
T. Yamaguchif;Yasuo KomodaP;H. Nakajima
本文首次报道了在人肝胞质组分中胆绿素还原酶的四种形式，包括两种胆绿素-MP还原酶和两种胆绿素-MA还原酶，分别命名为同工酶I和I1以及同工酶I11和IV。将胆绿素还原酶的四种形式纯化至均一。与粗品相比，比活性提高了7，80 ~ 15，000倍，回收率为8- 26%。纯化的酶是分子量约为21，000（同工酶I和11）和34，000（同工酶I11和IV）的单体。酶是严格的胆绿素特异性，并没有其他氧化还原酶的活动中检测到纯化的制剂。纯化的酶以NADPH和NADH为电子供体还原胆绿素。同工酶Ⅰ、Ⅱ、Ⅱ 1和Ⅳ对NADPH的表观K_i值分别为35.9、13.1、10.9和34.1 μ m，对NADH的表观K_i值分别为5.6、8.2、7.9和23.4 μ m。推测在胆绿素的胞内还原过程中，NADPH而不是NADH是生理电子供体。在NADPH体系中，胆绿素-MP同工酶Ⅰ和Ⅰ_1的表观K_（max）值为0.3 μ m，而胆绿素-MA同工酶Ⅰ_11和Ⅳ的表观K_（max）值分别为1.0 μ m和0.8 μ m。同工酶I和I1使用胆绿素-MP，-My和-MS作为底物，但不使用胆绿素Ma，同工酶I11和IV优选胆绿素-Ma作为四种胆绿素异构体中最有效的底物。当底物浓度超过34 p~+时，NADPH依赖性酶活性受到抑制。NADPH依赖酶活性，尤其是同工酶I11和IV，对SH试剂包括碘乙酰胺，对氯汞苯甲酸，N-乙基马来酰亚胺敏感。同工酶I和I1与NADPH反应的最适pH为8.2，而同工酶I11和IV的最适pH为7.4。同工酶I和11的总活性与同工酶I11和IV的总活性的比例在胎儿肝脏中比在成人肝脏中高得多。
This report describes for the first time the identification of four forms of biliverdin reductase including two biliverdin-MP reductases and two biliverdin-Ma reductases, designated isozymes I and I1 and isozymes I11 and IV, respectively, in human liver cytosolic fractions. The four forms of biliverdin reductase were purified to homogeneity. There was a 7,80&15,000-fold increase in specific activity when compared with the crude preparation, and the recovery was 8-26%. The purified enzymes were monomers with a molecular weight of about 21,000 (isozymes I and 11) and 34,000 (isozymes I11 and IV). The enzymes were strictly specific for biliverdin, and no other oxidoreductase activities were detected in the purified preparations. The purified enzymes used NADPH and NADH as electron donors for the reduction of biliverdin. The apparent K, values of isozymes I, 11,111, and IV for NADPH were 35.9, 13.1, 10.9, and 34.1 JIM, respectively, whereas those for NADH were 5.6, 8.2, 7.9, and 23.4 m~, respectively. It was assumed that NADPH rather than NADH was the physiological electron donor in the intracellular reduction of biliverdin. "he apparent K,,, value of isozymes I and I1 for biliverdin-MP in the NADPH system was 0.3 p~ whereas those of isozymes I11 and IV for biliverdin-Ma were 1.0 and 0.8 JIM, respectively. Isozymes I and I1 used biliverdin-MP, -My, and -MS as substrates but not biliverdinMa, and isozymes I11 and IV preferred biliverdin-Ma as the most effective substrate among the four biliverdin isomers. The NADPH-dependent enzyme activities were inhibited by substrate concentrations in excess of 34 p~. The NADPH-dependent enzyme activities, especially isozymes I11 and IV, were sensitive to SH reagents including iodoacetamide, p-chloromercuribenzoic acid, and N-ethylmaleimide. The optimum pH of the reaction with NADPH for isozymes I and I1 was 8.2 whereas that for isozymes I11 and IV was 7.4. The proportion of the total activity of isozymes I and 11 to that of isozymes I11 and IV was considerably higher in the fetal than in the adult liver.