溶组织内阿米巴钙结合蛋白5的克隆、纯化、结晶及初步晶体学研究
Cloning, purification, crystallization and preliminary crystallographic study of calcium-binding protein 5 from Entamoeba histolytica.
作者信息
Kumar Sanjeev, Zaidi Rana, Gourinath Samudrala
机构信息
School of Life Sciences, Jawaharlal Nehru University, New Delhi 110 067, India.
出版信息
Acta Crystallogr Sect F Struct Biol Cryst Commun. 2012 Dec 1;68(Pt 12):1542-4. doi: 10.1107/S1744309112044612. Epub 2012 Nov 19.
Abstract
Entamoeba histolytica is the causative agent of human amoebiasis. Phagocytosis is the major route of food intake by this parasite and is responsible for its virulence. Calcium and calcium-binding proteins play major roles in its phagocytosis. Calcium-binding protein 5 from E. histolytica (EhCaBP5) is a cytoplasmic protein; its expression is very sensitive to serum starvation and it seems to be involved in binding to myosin I. In this study, EhCaBP5 was cloned, expressed in Escherichia coli and purified using affinity and size-exclusion chromatography. The purified protein crystallized in space group C222 and the crystals diffracted to 2 Å resolution. The Matthews coefficient indicated the presence of one molecule in the asymmetric unit, with a VM of 2.35 Å3 Da(-1) and a solvent content of 47.7%.
摘要
溶组织内阿米巴是人类阿米巴病的病原体。吞噬作用是这种寄生虫摄取食物的主要途径，也是其毒力的来源。钙和钙结合蛋白在其吞噬作用中起主要作用。来自溶组织内阿米巴的钙结合蛋白5（EhCaBP5）是一种细胞质蛋白；其表达对血清饥饿非常敏感，似乎参与与肌球蛋白I的结合。在本研究中，克隆了EhCaBP5，在大肠杆菌中表达，并使用亲和色谱和尺寸排阻色谱进行纯化。纯化后的蛋白质在空间群C222中结晶，晶体衍射分辨率达到2 Å。马修斯系数表明不对称单位中存在一个分子，VM为2.35 Å3 Da-1，溶剂含量为47.7%。
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