Structure of a hyperthermophilic archaeal homing endonuclease, I-Tsp061I: Contribution of cross-domain polar networks to thermostability
Structure of a hyperthermophilic archaeal homing endonuclease, I-Tsp061I: Contribution of cross-domain polar networks to thermostability
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DOI：
10.1016/j.jmb.2006.09.066
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发表时间：
2007-01-12
影响因子：
5.6
通讯作者：
Tsuge, Hideaki
中科院分区：
文献类型：
--
作者：
Nakayama, Hitoshi;Shimamura, Tatsuro;Tsuge, Hideaki
从嗜热古菌Thermoproteus sp. IC-061的16 S rRNA基因（rDNA）内含子中分离得到一种新的LAGLIDADG型归巢核酸内切酶（HEase）I-TspO 61 I，并对其结构、催化性质和热稳定性进行了研究。发现I-Tsp 061 I是先前描述的I-PogI的HEase异构体，并且在已知的LAGLIDADG型HEase中表现出最高的热稳定性。使用多重同晶置换和异常散射方法在2.1 A分辨率下测定I-Tsp 061 I的晶体结构，结果表明，尽管I-TspO 61 I与其他已知的LAGLIDADG型HEases之间的序列相似性很小，但总体折叠与其他已知的LAGLIDADG型HEases相似。然而，I-Tsp 061 I包含重要的跨域极性网络，与其嗜温对应物不同。值得注意的是，极性网络Tyr 6-Asp 104-His 180 - 107 O-HOH 12 - 104 O-Asn 177存在于包含LAGLIDADG催化基序和GxxxG疏水螺旋束基序的两个堆积的α-螺旋上。另一个重要的结构特征是跨越N和C-末端结构域界面的盐桥网络Asp 29-Arg 31-GIu 182，其似乎有助于结构域/结构域包装的稳定性。在这些结构分析和广泛的突变研究的基础上，我们得出结论，这种跨域极性网络在稳定催化中心和域包装中起着关键作用，是T-Tsp 061 I的超热稳定性的基础。(c)2006爱思唯尔有限公司出版
A novel LAGLIDADG-type homing endonuclease (HEase), I-TspO61I, from the hyperthermophilic archaeon Thermoproteus sp. IC-061 16 S rRNA gene (rDNA) intron was characterized with respect to its structure, catalytic properties and thermostability. It was found that I-Tsp061I is a HEase isoschizomer of the previously described I-PogI and exhibits the highest thermostability among the known LAGLIDADG-type HEases. Determination of the crystal structure of I-Tsp061I at 2.1 A resolution using the multiple isomorphous replacement and anomalous scattering method revealed that the overall fold is similar to that of other known LAGLIDADG-type HEases, despite little sequence similarity between I-TspO61I and those HEases. However, I-Tsp061I contains important cross-domain polar networks, unlike its mesophilic counterparts. Notably, the polar network Tyr6-Asp104-His180-107O-HOH12-104O-Asn177 exists across the two packed a-helices containing both the LAGLIDADG catalytic motif and the GxxxG hydrophobic helix bundle motif. Another important structural feature is the salt-bridge network Asp29-Arg31-GIu182 across N and C-terminal domain interface, which appears to contribute to the stability of the domain/domain packing. On the basis of these structural analyses and extensive mutational studies, we conclude that such cross-domain polar networks play key roles in stabilizing the catalytic center and domain packing, and underlie the hyperthermostability of T-Tsp061I. (c) 2006 Published by Elsevier Ltd.