DNA nicking by HinP1I endonuclease: bending, base flipping and minor groove expansion
DNA nicking by HinP1I endonuclease: bending, base flipping and minor groove expansion
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DOI：
10.1093/nar/gkj484
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发表时间：
2006-01-01
影响因子：
14.9
通讯作者：
Cheng, XD
中科院分区：
文献类型：
--
作者：
Horton, JR;Zhang, X;Cheng, XD
HINP1 I识别并切割DNA中的palindromic四核苷酸序列g向下箭头CGC。我们报告了HINP1 I-DNA复合物的三个结构：在没有金属离子（二进制复合物）的情况下，并且在MG2+（反应后复合物）的情况下存在Ca2+（反应性复合物）。 HINP1 I形成了一个背对背二聚体，具有两个活动位点和两个DNA双链体，这些二聚体在二聚体的外表面上，彼此面向二聚体。 10 bp DNA双链体经历蛋白质引起的扭曲，表现出A-，B和Z形成的特征：一侧弯曲（通过将苯丙氨酸侧链插入到主要的凹槽中），底座在识别的另一侧翻转站点（通过扩大本地基对的步长上升距离至z形式）和识别位点的两个中央C：G基础对之间的局部A形式构象（通过在小凹槽中结合N末端螺旋）。在反应后和反应性复合物中，在活动位点发现了两种金属（Ca2+或Mg2+）。该酶似乎依次裂解DNA，水解第一个DNA链，如在晶体状态下反应性复合物中所见，然后在溶液中的观察中支持了另一个DNA链，而在溶液中，一个成形的DNA中间体在线性化前会积聚。
HinP1 I recognizes and cleaves the palindromic tetranucleotide sequence G down arrow CGC in DNA. We report three structures of HinP1 I-DNA complexes: in the presence of Ca2+ (pre-reactive complex), in the absence of metal ion (binary complex) and in the presence of Mg2+ (post-reactive complex). HinP1 I forms a back-to-back dimer with two active sites and two DNA duplexes bound on the outer surfaces of the dimer facing away from each other. The 10 bp DNA duplexes undergo protein-induced distortions exhibiting features of A-, B- and Z-conformations: bending on one side (by intercalation of a phenylalanine side chain into the major groove), base flipping on the other side of the recognition site (by expanding the step rise distance of the local base pair to Z-form) and a local A-form conformation between the two central C:G base pairs of the recognition site (by binding of the N-terminal helix in the minor groove). In the pre- and post-reactive complexes, two metals (Ca2+ or Mg2+) are found in the active site. The enzyme appears to cleave DNA sequentially, hydrolyzing first one DNA strand, as seen in the post-reactive complex in the crystalline state, and then the other, as supported by the observation that, in solution, a nicked DNA intermediate accumulates before linearization.