The structure of a bacterial L-amino acid oxidase from Rhodococcus opacus gives new evidence for the hydride mechanism for dehydrogenation
The structure of a bacterial L-amino acid oxidase from Rhodococcus opacus gives new evidence for the hydride mechanism for dehydrogenation
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DOI：
10.1016/j.jmb.2006.11.071
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发表时间：
2007-03-16
影响因子：
5.6
通讯作者：
Schomburg, Dietmar
中科院分区：
文献类型：
--
作者：
Faust, Annette;Niefind, Karsten;Schomburg, Dietmar
根据辅助因子结合的高度保守序列基序，来自不透明红球菌的l -氨基酸氧化酶（roLAAO）被归类为黄素依赖性氧化还原酶GR(2)-家族的成员。同型二聚体酶的单体由三个定义明确的结构域组成：与整个gr2家族的一般拓扑结构相对应的fad结合结构域；一个与蛇毒LAAO结构几乎相同的底物结合结构域和一个螺旋结构域，专门负责酶的不寻常的二聚化模式，迄今未在该家族的其他成员中发现。我们描述了蛋白质和辅因子的二元配合物以及蛋白质、辅因子和配体的三元配合物的高分辨率结构。这种结构加上蛇毒LAAO和酵母和猪肾DAAO的结构知识，可以更深入地了解这类酶的反应机制的不同步骤。氢化物转移是脱氢反应的主要机理。这种机制似乎是不常见的，在某种意义上，化学转化可以有效地进行没有氨基酸官能团的参与。活性位点上的大多数基团参与底物识别、结合和固定，即它们指导相互作用轨道的轨迹。在这种催化模式中，轨道转向/相互作用是化学步骤的主要因素。D和l -氨基酸氧化酶的两个底物结合位点之间存在镜像对称关系，这有利于对映体的选择性，同时保持活性位点残基的共同排列。这些结果对黄酮类蛋白的机制具有普遍的相关性，并导致在L和d氨基酸氧化酶的机制中提出一个共同的脱氢步骤。(c) 2006 Elsevier Ltd.版权所有。
L-Amino acid oxidase from Rhodococcus opacus (roLAAO) is classified as a member of the GR(2)-family of flavin-dependent oxidoreductases according to a highly conserved sequence motif for the cofactor binding. The monomer of the homodimeric enzyme consists of three well-defined domains: the FAD-binding domain corresponding to a general topology throughout the whole GR2-family; a substrate-binding domain with almost the same topology as the snake venom LAAO and a helical domain exclusively responsible for the unusual dimerisation mode of the enzyme and not found in other members of the family so far.We describe here high-resolution structures of the binary complex of protein and cofactor as well as the ternary complexes of protein, cofactor and ligands. This structures in addition to the structural knowledge of snake venom LAAO and DAAO from yeast and pig kidney permit more insight into different steps in the reaction mechanism of this class of enzymes. There is strong evidence for hydride transfer as the mechanism of dehydrogenation. This mechanism appears to be uncommon in a sense that the chemical transformation can proceed efficiently without the involvement of amino acid functional groups. Most groups present at the active site are involved in substrate recognition, binding and fixation, i.e. they direct the trajectory of the interacting orbitals. In this mode of catalysis orbital steering/interactions are the predominant factors for the chemical step(s). A mirror-symmetrical relationship between the two substrate-binding sites Of D and L-amino acid oxidases is observed which facilitates enantiomeric selectivity while preserving a common arrangement of the residues in the active site. These results are of general relevance for the mechanism of flavoproteins and lead to the proposal of a common dehydrogenation step in the mechanism for L and D-amino acid oxidases. (c) 2006 Elsevier Ltd. All rights reserved.