Inoue Kengo, Ashikawa Yuji, Umeda Takashi, Abo Mitsuru, Katsuki Junichi, Usami Yusuke, Noguchi Haruko, Fujimoto Zui, Terada Tohru, Yamane Hisakazu, Nojiri Hideaki
Biotechnology Research Center, The University of Tokyo, Tokyo, Japan.
J Mol Biol. 2009 Sep 18;392(2):436-51. doi: 10.1016/j.jmb.2009.07.029. Epub 2009 Jul 17.
Carbazole 1,9a-dioxygenase (CARDO) consists of terminal oxygenase (Oxy), ferredoxin (Fd), and ferredoxin reductase (Red) components and is a member of the Rieske nonheme iron oxygenases. Rieske nonheme iron oxygenases are divided into five subclasses (IA, IB, IIA, IIB, and III) based on the number of constituents and the nature of their redox centers. Each component of a class IIB CARDO from Nocardioides aromaticivorans IC177 was purified, and the interchangeability of the electron transfer reactions with each component from the class III CARDOs was investigated. Despite the fact that the Fds of both classes are Rieske-type, strict specificities between the Oxy and Fd components were observed. On the other hand, the Fd and Red components were interchangeable, even though the Red components differ in cofactor composition; the class IIB Red contains flavin-adenine-dinucleotide (FAD)- and NADH-binding domains, whereas the class III Red has a chloroplast-type [2Fe-2S] cluster in addition to the FAD- and NADH-binding domains. The crystal structures of the class IIB Oxy and Fd components were compared to the previously reported Fd:Oxy complex structure of class III CARDO. This comparison suggested residues in common between class IIB and class III CARDOs that are important for interactions between Fd and Oxy. In the class IIB CARDOs, these included His75 and Glu71 in Fd and Lys20 and Glu357 in Oxy for electrostatic interactions, and Phe74 and Pro90 in Fd and Trp21, Leu359, and Val367 in Oxy for hydrophobic interactions. The residues that formed the interacting surface but were not conserved between classes were thought to be necessary to form the appropriate geometry and to determine electron transfer specificity between Fd and Oxy.
咔唑1,9a -双加氧酶（CARDO）由末端加氧酶（Oxy）、铁氧化还原蛋白（Fd）和铁氧化还原蛋白还原酶（Red）组成，是 Rieske 非血红素铁加氧酶家族的一员。Rieske 非血红素铁加氧酶根据其组成成分的数量及其氧化还原中心的性质分为五个亚类（IA、IB、IIA、IIB 和 III）。对来自食芳烃诺卡氏菌IC177的IIB类CARDO的每个组分进行了纯化，并研究了其与III类CARDO的各组分之间电子转移反应的互换性。尽管两类的Fd都是 Rieske 型，但观察到Oxy和Fd组分之间存在严格的特异性。另一方面，尽管Red组分的辅因子组成不同，但Fd和Red组分是可互换的；IIB类Red含有黄素腺嘌呤二核苷酸（FAD）和NADH结合结构域，而III类Red除了FAD和NADH结合结构域外，还有一个叶绿体型[2Fe - 2S]簇。将IIB类Oxy和Fd组分的晶体结构与先前报道的III类CARDO的Fd:Oxy复合结构进行了比较。这种比较表明，IIB类和III类CARDO之间存在共同的残基，这些残基对于Fd和Oxy之间的相互作用很重要。在IIB类CARDO中，这些残基包括Fd中的His75和Glu71以及Oxy中的Lys20和Glu357用于静电相互作用，Fd中的Phe74和Pro90以及Oxy中的Trp21、Leu359和Val367用于疏水相互作用。形成相互作用表面但在不同类别之间不保守的残基被认为是形成合适几何结构以及确定Fd和Oxy之间电子转移特异性所必需的。
