Structural Basis of Biological Nitrile Reduction
Structural Basis of Biological Nitrile Reduction
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DOI：
10.1074/jbc.m112.388538
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发表时间：
2012-08-31
影响因子：
4.8
通讯作者：
Swairjo, Manal A.
中科院分区：
文献类型：
--
作者：
Chikwana, Vimbai M.;Stec, Boguslaw;Swairjo, Manal A.
酶QUF催化7-氰基-7-去氮鸟嘌呤(preQ(0))的氰基还原为7-氨甲基-7-去氮鸟嘌呤(preQ(1))，这是生物学上已知的唯一的氰基还原反应。我们描述了枯草芽孢杆菌QueF的两种晶体结构，一种是野生型酶，与底物preQ(0)形成络合物，以共价硫酰亚胺的形式被捕获，可能是反应中的中间体；第二种是C55A突变体，与底物preQ(0)以非共价结合。QueF酶形成一个由两个面对面的五聚体亚基组成的不对称隧道折叠同源十聚体，在亚基间的界面上有10个活性位点。在这两种结构中，preQ(0)分子结合在8个位置，在野生型酶中，它与催化残基Cys-55形成硫代亚胺共价键。结构和瞬时动力学数据都表明，前Q(0)结合而不是硫代亚胺的形成导致了活性中心的大的构象变化和关闭。基于这些数据，我们提出了一种激活Cys-55亲核试剂和随后的氢化物转移的机制。
The enzyme QueF catalyzes the reduction of the nitrile group of 7-cyano-7-deazaguanine (preQ(0)) to 7-aminomethyl-7-deazaguanine (preQ(1)), the only nitrile reduction reaction known in biology. We describe here two crystal structures of Bacillus subtilis QueF, one of the wild-type enzyme in complex with the substrate preQ(0), trapped as a covalent thioimide, a putative intermediate in the reaction, and the second of the C55A mutant in complex with the substrate preQ(0) bound noncovalently. The QueF enzyme forms an asymmetric tunnel-fold homodecamer of two head-to-head facing pentameric subunits, harboring 10 active sites at the intersubunit interfaces. In both structures, a preQ(0) molecule is bound at eight sites, and in the wild-type enzyme, it forms a thioimide covalent linkage to the catalytic residue Cys-55. Both structural and transient kinetic data show that preQ(0) binding, not thioimide formation, induces a large conformational change in and closure of the active site. Based on these data, we propose a mechanism for the activation of the Cys-55 nucleophile and subsequent hydride transfer.