采用X射线衍射解析人鸟嘌呤核苷酸解离刺激因子（RalGDS）Ras结合结构域的空间结构

Ras结合结构域（RBD）是鸟嘌呤核苷酸解离刺激因子（Ral GDS）家族成员C-端的高保守区,通过它连接Ras和Ras相关蛋白。利用Red Wings和SGC-1 screens相关的悬滴法设盘结晶,按体积比1∶1加蛋白液到含1∶100胞内蛋白酶Glu-C（w/w）的结晶溶液（2 mol/L（NH4）2SO4,0.2 mol/L Na Ac,0.1 mol/L HEPES,5%MPD,p H 7.5）中,晶体3天长成可组装大小。利用X-射线晶体衍射技术解析了人Ral GDS的Ras结合域（Ral GDS-RBD）的晶体结构,对比鼠和人Ral GDS-RBD,主要是Ras结合区的C-端不同。人...

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Bibliographic Details
Published in分析化学 Vol. 43; no. 6; pp. 893 - 898
Main Author 黄雪英 雷明 沈阳 张巍 刘艳丽 刘珂 赵浩斌 祁超
Format Journal Article
LanguageChinese
Published 2015
Subjects
GDS
Ras结合结构域（RBD）
Ras蛋白
鸟嘌呤核苷酸解离刺激因子（Ral
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Summary:Ras结合结构域（RBD）是鸟嘌呤核苷酸解离刺激因子（Ral GDS）家族成员C-端的高保守区,通过它连接Ras和Ras相关蛋白。利用Red Wings和SGC-1 screens相关的悬滴法设盘结晶,按体积比1∶1加蛋白液到含1∶100胞内蛋白酶Glu-C（w/w）的结晶溶液（2 mol/L（NH4）2SO4,0.2 mol/L Na Ac,0.1 mol/L HEPES,5%MPD,p H 7.5）中,晶体3天长成可组装大小。利用X-射线晶体衍射技术解析了人Ral GDS的Ras结合域（Ral GDS-RBD）的晶体结构,对比鼠和人Ral GDS-RBD,主要是Ras结合区的C-端不同。人Ral GDS-RBD通过Glu838和Glu840在Ral GDS和Ras蛋白间形成氢键,而在同一位点,鼠Ral GDS-RBD通过Asp820和Asp822形成氢键。人Ral GDS-RBD结构中含ββαββαβ-型三维结构的泛素样构象,一个单体的C-端残基与相邻单体的β折叠形成平行βββββ结构。
Bibliography:HUANG Xue-Ying , LEI Ming, SHEN Yang, ZHANG Wei, LIU Yan-Li, LIU Ke, ZHAO Hao-Bin, QI Chao l （ College of Chemistry, Central China Normal University, Wuhan 430079, China） 2 （ Hubei Key Laboratory of Genetic Regulation and Integrative Biology, College of Life Sciences, Central China Normal University, Wuhan 430079, China） 3（Structural Genomics Consortium, University of Toronto, 101 College St., Toronto, Ontario, Canada M5G 1LT）
22-1125/O6
The Ras binding domain （RBD） is a highly conserved domain in the C-terminal region of Ral guanine nucleotide dissociation stimulator （RalGDS） , which functions as cross-linking domain between Ras and Ras-related proteins. Crystallization was setup using in situ proteolysis method of sitting drops with Red Wings and SGC-I screens. Crystals suitable for X-ray diffraction analysis were obtained by mixing equal volumes of the protein solution and the reservoir solution [ 2 mol/L （NH4 ）2SO4, 0. 2 mol/L NaAc, 0.1 mol/L N-hydroxyethyl piperazine ethanesulfonic acid （ HEPES）, 5 % 1,3-m
ISSN:0253-3820