The nuclease A-inhibitor complex is characterized by a novel metal ion bridge
The nuclease A-inhibitor complex is characterized by a novel metal ion bridge
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DOI：
10.1074/jbc.m605986200
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发表时间：
2007-02-23
影响因子：
4.8
通讯作者：
Pedersen, Lars C.
中科院分区：
文献类型：
--
作者：
Ghosh, Mahua;Meiss, Gregor;Pedersen, Lars C.
非特异性，细胞外核酸酶最近受到了越来越多的关注，因为这些酶可以通过克服宿主中性粒细胞防御系统在感染性中发挥关键作用。蓝藻核酸酶NucA是β β α Me超家族的成员，其活性受特异性核酸酶抑制剂NuiA控制。在这里，我们报告了NucA-NuiA复合物的2.3埃分辨率晶体结构，表明NuiA对NucA的抑制涉及一个不寻常的二价金属离子桥，该桥连接核酸酶及其抑制剂。C-末端Thr-135（NuiA）羟基氧直接与核酸酶活性位点的催化Mg 2+配位，并且Glu-24（NuiA）也延伸到活性位点中，模拟易断裂的磷酸盐的电荷。NuiA残基Asp-75和Trp-76形成第二相互作用位点，有助于相互作用的强度和特异性。晶体学定义的接口被证明是一致的研究结果，使用定点NuiA突变体。这种抑制模式与用DNA酶大肠杆菌素E7/E9和已研究的其他核酸酶抑制剂复合物观察到的外位点抑制机制显著不同。该复合物的结构为开发相关非特异性核酸酶的抑制剂提供了有价值的见解，所述非特异性核酸酶共享DRGH活性位点基序，如肺炎链球菌核酸酶EndA，其介导该病原体的感染性，以及线粒体EndoG，其参与重组和凋亡。
Nonspecific, extracellular nucleases have received enhanced attention recently as a consequence of the critical role that these enzymes can play in infectivity by overcoming the host neutrophil defense system. The activity of the cyanobacterial nuclease NucA, a member of the beta beta alpha Me superfamily, is controlled by the specific nuclease inhibitor, NuiA. Here we report the 2.3-angstrom resolution crystal structure of the NucA-NuiA complex, showing that NucA inhibition by NuiA involves an unusual divalent metal ion bridge that connects the nuclease with its inhibitor. The C-terminal Thr-135(NuiA) hydroxyl oxygen is directly coordinated with the catalytic Mg2+ of the nuclease active site, and Glu-24(NuiA) also extends into the active site, mimicking the charge of a scissile phosphate. NuiA residues Asp-75 and Trp-76 form a second interaction site, contributing to the strength and specificity of the interaction. The crystallographically defined interface is shown to be consistent with results of studies using site-directed NuiA mutants. This mode of inhibition differs dramatically from the exosite mechanism of inhibition seen with the DNase colicins E7/E9 and from other nuclease-inhibitor complexes that have been studied. The structure of this complex provides valuable insights for the development of inhibitors for related nonspecific nucleases that share the DRGH active site motif such as the Streptococcus pneumoniae nuclease EndA, which mediates infectivity of this pathogen, and mitochondrial EndoG, which is involved in recombination and apoptosis.