Unveiling the molecular mechanism of a conjugative relaxase: The structure of TrwC complexed with a 27-mer DNA comprising the recognition hairpin and the cleavage site
Unveiling the molecular mechanism of a conjugative relaxase: The structure of TrwC complexed with a 27-mer DNA comprising the recognition hairpin and the cleavage site
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DOI：
10.1016/j.jmb.2006.02.018
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发表时间：
2006-05-05
影响因子：
5.6
通讯作者：
de la Cruz, F
中科院分区：
文献类型：
--
作者：
Boer, R;Russi, S;de la Cruz, F
TrwC是一种DNA链转移酶，催化接合DNA转移的初始和最终阶段。我们已经解决了与27个碱基长的DNA寡核苷酸，其中包含识别发夹和易裂磷酸盐的复杂的N-末端松弛酶结构域的TrwC的晶体结构。此外，一系列的三元结构的蛋白质-DNA复合物与不同的二价阳离子的活性位点已被解决。系统的异常差异分析使我们能够明确地确定金属结合的性质。Zn ~（2+）、Ni ~（2+）和Cu ~（2+）与组氨酸三联体金属结合位点结合。不同复合物的结构的比较表明，两种途径的DNA退出的活性口袋。它们可能用于接合DNA加工反应的不同步骤。结构信息使我们能够提出（i）一种酶机制，其中易分裂的磷酸盐被金属离子极化，促进催化酪氨酸的亲核攻击，以及（ii）结合DNA加工过程中可能的事件序列，解释了生物学功能的松弛酶。(c)2006爱思唯尔有限公司保留所有权利。
TrwC is a DNA strand transferase that catalyzes the initial and final stages of conjugative DNA transfer. We have solved the crystal structure of the N-terminal relaxase domain of TrwC in complex with a 27 base-long DNA oligonucleotide that contains both the recognition hairpin and the scissile phosphate. In addition, a series of ternary structures of protein-DNA complexes with different divalent cations at the active site have been solved. Systematic anomalous difference analysis allowed us to determine unambiguously the nature of the metal bound. Zn2+, Ni2+ and Cu2+ were found to bind the histidine-triad metal binding site. Comparison of the structures of the different complexes suggests two pathways for the DNA to exit the active pocket. They are probably used at different steps of the conjugative DNA-processing reaction. The structural information allows us to propose (i) an enzyme mechanism where the scissile phosphate is polarized by the metal ion facilitating the nucleophilic attack of the catalytic tyrosine, and (ii) a probable sequence of events during conjugative DNA processing that explains the biological function of the relaxase. (c) 2006 Elsevier Ltd. All rights reserved.