Structural studies of E-coli topoisomerase III-DNA complexes reveal a novel type IA topoisomerase-DNA conformational intermediate
Structural studies of E-coli topoisomerase III-DNA complexes reveal a novel type IA topoisomerase-DNA conformational intermediate
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DOI：
10.1016/j.jmb.2007.01.065
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发表时间：
2007-04-20
影响因子：
5.6
通讯作者：
Mondragon, Alfonso
中科院分区：
文献类型：
--
作者：
Changela, Anita;DiGate, Russell J.;Mondragon, Alfonso
大肠杆菌DNA拓扑异构酶III属于DNA拓扑异构酶的IA型家族，其通过51磷酸酪氨酸中间体瞬时切割单链DNA（ssDNA）。我们已经解决了野生型E的晶体结构。coli拓扑异构酶III在三种不同pH环境中与八个碱基的ssDNA分子结合。结构显示酶在与DNA结合时处于三种不同的构象状态。一种构象类似于先前观察到的与DNA结合的拓扑异构酶III的无催化活性的突变体，其中DNA结合重新排列催化残基以形成功能活性位点。另一种构象代表了一种新的中间体，其中DNA沿着ssDNA结合沟结合，但不进入活性位点，其保持在催化失活的封闭状态。第三种构象显示了一种中间状态，在这种状态下，酶仍然处于闭合状态，但ssDNA开始侵入活性位点。对于第一次，在催化酪氨酸和ssDNA底物的存在下的活性位点区域被揭示为IA型DNA拓扑异构酶，虽然没有证据ssDNA裂解。各种构象状态的比较分析表明，底物结合后的酶进行的结构域运动的序列。(c)2007爱思唯尔有限公司保留所有权利。
Escherichia coli DNA topoisomerase III belongs to the type IA family of DNA topoisomerases, which transiently cleave single-stranded DNA (ssDNA) via a 51 phosphotyrosine intermediate. We have solved crystal structures of wild-type E. coli topoisomerase III bound to an eight-base ssDNA molecule in three different pH environments. The structures reveal the enzyme in three distinct conformational states while bound to DNA. One conformation resembles the one observed previously with a DNA-bound, catalytically inactive mutant of topoisomerase III where DNA binding realigns catalytic residues to form a functional active site. Another conformation represents a novel intermediate in which DNA is bound along the ssDNA-binding groove but does not enter the active site, which remains in a catalytically inactive, closed state. A third conformation shows an intermediate state where the enzyme is still in a closed state, but the ssDNA is starting to invade the active site. For the first time, the active site region in the presence of both the catalytic tyrosine and ssDNA substrate is revealed for a type IA DNA topoisomerase, although there is no evidence of ssDNA cleavage. Comparative analysis of the various conformational states suggests a sequence of domain movements undertaken by the enzyme upon substrate binding. (c) 2007 Elsevier Ltd. All rights reserved.