Basic and applied features of multicopper oxidases, CueO, bilirubin oxidase, and laccase
Basic and applied features of multicopper oxidases, CueO, bilirubin oxidase, and laccase
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DOI：
10.1002/tcr.20125
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发表时间：
2007-01-01
期刊：
影响因子：
6.6
通讯作者：
Kataoka, Kunishige
中科院分区：
文献类型：
--
作者：
Sakurai, Takeshi;Kataoka, Kunishige
多铜氧化酶(MCO)如CueO、胆红素氧化酶和漆酶，在一个由三个结构域组成的蛋白质分子中含有四个铜中心，即I型铜、II型铜和一对III型铜，这些结构域与只含有I型铜的铜还蛋白具有同源结构。I型铜介导了底物与由II型铜和一对III型铜形成的三核铜中心之间的电子转移，最终的电子受体O-2被转化为H2O，而不释放活性氧物种。在此过程中，0(2)被漆酶和胆红素氧化酶等MCOs还原，检测到反应中间体II在三核铜中心可能具有双OH-桥联结构。通过漆酶在混合价状态下的反应检测到前一反应中间体I，在混合价状态下，I型铜被完全还原，三核铜中心被完全还原，胆红素氧化酶中I型铜位置的Cys->；Ser突变体与CueO反应。位于三核铜中心附近的酸性氨基酸残基被证明是这些反应中间产物的质子供体。MCO对有机底物的底物专一性是底物结合部位的形状和底物与与I型铜配位的His残基附近氨基酸的特异性相互作用的综合作用。相反，亚铜氧化酶CueO的底物专一性是通过覆盖铜(I)结合部位的片段来阻碍有机底物的访问而产生的。截短跨螺旋5-7的片段大大降低了CueO对铜(I)的专一性，显著提高了对有机底物的低氧化活性，表明蛋白质工程修饰MCO底物专一性是成功的。(C)2007年日本化学期刊论坛和威利期刊公司。
Multicopper oxidases (MCOs) such as CueO, bilirubin oxidase, and laccase contain four Cu centers, type 1 Cu, type II Cu, and a pair of type III Cu's in a protein molecule consisting of three domains with homologous structure to cupredoxin containing only type I Cu. Type I Cu mediates electron transfer between the substrate and the trinuclear Cu center formed by a type II Cu and a pair of type III Cu's, where the final electron acceptor O-2 is converted to H2O without releasing activated oxygen species. During the process; 0(2) is reduced by MCOs such as lacquer laccase and bilirubin oxidase; the reaction intermediate II with a possible doubly OH--bridged structure in the trinuclear Cu center has been detected. The preceding reaction intermediate I has been detected by the reaction of the lacquer laccase in a mixed valence state, at which type I Cu was cuprous and the trinuclear Cu center was fully reduced, and by the reaction of the Cys -> Ser mutant for the type I Cu site in bilirubin oxidase and CueO. An acidic amino acid residue located adjacent to the trinuclear Cu center was proved to function as a proton donor to these reaction intermediates. The substrate specificity of MCO for organic substrates is produced by the integrated effects of the shape of the substrate-binding site and the specific interaction of the substrate with the amino acid located adjacent to the His residue coordinating to the type I Cu. In contrast, the substrate specificity of the cuprous oxidase, CueO, is produced by the segment covering the Cu(I)-binding site so as to obstruct the access of organic substrates. Truncating the segment spanning helix 5 to helix 7 greatly reduced the specificity of CueO for Cu(I) and prominently enhanced the low oxidizing activity for the organic substrates, indicating the success of protein engineering to modify the substrate specificity of MCO. (c) 2007 The Japan Chemical Journal Forum and Wiley Periodicals, Inc.