STRUCTURE OF THE DNA-ECO RI ENDONUCLEASE RECOGNITION COMPLEX AT 3 A RESOLUTION
STRUCTURE OF THE DNA-ECO RI ENDONUCLEASE RECOGNITION COMPLEX AT 3 A RESOLUTION
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DOI：
10.1126/science.3024321
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发表时间：
1986-12-19
期刊：
影响因子：
56.9
通讯作者：
ROSENBERG, JM
中科院分区：
文献类型：
--
作者：
MCCLARIN, JA;FREDERICK, CA;ROSENBERG, JM
Eco RI内切酶与同源寡核苷酸TCGCGAATTCGCG之间的复合物的晶体结构为序列特异性DNA-蛋白质相互作用的结构基础提供了详细的例子。用具有铂同晶导数的迭代单同晶置换(ISIR)方法确定了结构，分辨率为3？该复合体具有双重对称性。核酸内切酶的每个亚基都被组织成一个α/β结构域，它由一个五链的β片层、α螺旋和一个被称为“臂”的延伸部分组成。大的β片断由反平行和平行基序组成，这些基序构成了分别负责DNA链断裂和序列特异性识别的环和α螺旋的基础。DNA裂解位点位于一个裂隙中，该裂隙将DNA主干结合在剪切键附近。序列特异性由来自α螺旋识别模块的12个氢键介导。Arg200与鸟嘌呤形成两个氢键，而Glu144和Arg145与邻近的腺嘌呤残基形成四个氢键。这些相互作用将Eco RI六核苷酸GAATTC与所有其他六核苷酸区分开来，因为任何碱基取代都需要至少打断其中一个氢键。
The crystal structure of the complex between Eco RI endonuclease and the cognate oligonucleotide TCGCGAATTCGCG provides a detailed example of the structural basis of sequence-specific DNA-protein interactions. The structure was determined, to 3 Å resolution, by the ISIR (iterative single isomorphous replacement) method with a platinum isomorphous derivative. The complex has twofold symmetry. Each subunit of the endonuclease is organized into an α/β domain consisting a five-stranded β sheet, α helices, and an extension, called the "arm," which wraps around the DNA. The large β sheet consists of antiparallel and parallel motifs that form the foundations for the loops and α helices responsible for DNA strand scission and sequence-specific recognition, respectively. The DNA cleavage site is located in a cleft that binds the DNA backbone in the vicinity of the scissile bond. Sequence specificity is mediated by 12 hydrogen bonds originating from α helical recognition modules. Arg200forms two hydrogen bonds with guanine while Glu144and Arg145form four hydrogen bonds to adjacent adenine residues. These interactions discriminate the Eco RI hexanucleotide GAATTC from all other hexanucleotides because any base substitution would require rupture of at least one of these hydrogen bonds.