Crystallographic analysis of the Arabidopsis thaliana BAG5-calmodulin protein complex
Crystallographic analysis of the Arabidopsis thaliana BAG5-calmodulin protein complex
复制标题
拟南芥 BAG5-钙调蛋白复合物的晶体分析。
DOI：
10.1107/s2053230x15005956
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发表时间：
2015-07-01
影响因子：
0.9
通讯作者：
Yang, Xue
中科院分区：
文献类型：
--
作者：
Cui, Boyang;Fang, Shasha;Yang, Xue
拟南芥BAG5(AtBAG5)属于植物包(Bcl2相关基因)家族，具有从生长发育到非生物胁迫和衰老的多种功能。BAG家族成员可以通过其进化保守的BAG结构域与Hsp70 ATPase结构域结合，作为热休克蛋白70(Hsp70)的核苷酸交换因子，从而可能参与植物伴侣介导的蛋白质折叠的调节。与其他家族成员不同的是，AtBAG5在BAG结构域附近存在一个独特的IQ基序；这个基序是钙调蛋白(CaM)结合的特异性基序，表明它在植物钙信号通路中具有潜在的作用。为了更好地理解IQ基序介导的AtBAG5和CaM之间的相互作用，我们分别表达和纯化了这两种蛋白，然后将它们共结晶在一起。在0.1 M Tris-HCl pH 8.5，2.5 M硫酸铵的条件下，用坐滴气相扩散法生长了衍射级晶体。晶体属P2(1)2(1)2(1)空间群，晶胞参数a=64.56，b=74.89，c=117.09 ä。X射线衍射数据是用同步辐射从单晶记录到2.5 ä的分辨率。假定不对称单元中存在两个分子，则计算出马修斯系数为2.44 ä(3) Da(-1)，对应的溶剂含量约为50%。
Arabidopsis thaliana BAG5 (AtBAG5) belongs to the plant BAG (Bcl-2-associated athanogene) family that performs diverse functions ranging from growth and development to abiotic stress and senescence. BAG family members can act as nucleotide-exchange factors for heat-shock protein 70 (Hsp70) through binding of their evolutionarily conserved BAG domains to the Hsp70 ATPase domain, and thus may be involved in the regulation of chaperone-mediated protein folding in plants. AtBAG5 is distinguished from other family members by the presence of a unique IQ motif adjacent to the BAG domain; this motif is specific for calmodulin (CaM) binding, indicating a potential role in the plant calcium signalling pathway. To provide a better understanding of the IQ motif-mediated interaction between AtBAG5 and CaM, the two proteins were expressed and purified separately and then co-crystallized together. Diffraction-quality crystals of the complex were grown using the sitting-drop vapour-diffusion technique from a condition consisting of 0.1 M Tris-HCl pH 8.5, 2.5 M ammonium sulfate. The crystals belonged to space group P2(1)2(1)2(1), with unit-cell parameters a = 64.56, b = 74.89, c = 117.09 Å. X-ray diffraction data were recorded to a resolution of 2.5 Å from a single crystal using synchrotron radiation. Assuming the presence of two molecules in the asymmetric unit, a Matthews coefficient of 2.44 Å(3) Da(-1) was calculated, corresponding to a solvent content of approximately 50%.