大肠杆菌胞嘧碱通透酶CodB结构和功能的研究

The Nucleobase/Cation Symporter-1 (NCS1) Family consists of over 1000 currently sequenced proteins, including Gram-negative and Gram-positive bacteria, archaea, yeast, fungi and plants. Among the proteins identified, the bacterial and yeast proteins could not cluster closely on the NCS1 family phylogenetic tree with divergent differentiation .The nucleobase/cation symporter-2 proteins，are responsible for the uptake of nucleobases in all kingdoms of life. Apart from thatNCS1 proteins play vital roles in elimination of metabolic waste products in certain species. Despite their increasing biological importance, studying NCS1 family members has been an on-going challenge due to the vast lack of knowledge in three-dimensional structure and functional characterization of the NCS1. Besides Mhp1, to decipher the other proteins' detailed structural and to elucidate their molecular information remain outstanding task.Here we choose the Escherichia coli cytosine permease CodB as main research object. It is important for the study of its three-dimensional crystal structure to enable comprehensive understanding of the molecular mechanism of NCS1 family protein transporter substrates into cells.Moreover, through detailed comparison to the known structure of NCS2 family proteins, we tend to explore common structural features and molecular between these transporter substrates and further enhance the understanding of the transporter..

核糖碱基/阳离子同向转运蛋白-1（NCS1）家族涵盖了来自革兰氏阴性菌、革兰氏阳性菌、古生菌、酵母、真菌、植物等超过1000个蛋白成员。其中来自细菌和真菌的成员之间存在着明显的差异，在进化树上不能得到严密的聚类。NCS1家族蛋白在不同种类的生物中都负责摄取外源性核酸碱基。除此之外，该家族蛋白还在某些生物体内履行着转运代谢废物排出细胞的职责。尽管这类家族蛋白的生物学意义如此重要，除Mhp1以外，尚无任何一个细菌、真菌或者哺乳动物的NCS1家族蛋白的三维晶体结构被解析出来，对其功能特征的描述更是无从谈起。我们选择大肠杆菌胞嘧啶通透酶CodB为主要研究对象，希望通过对其三维晶体结构进行解析来了解NCS1家族蛋白转运底物进入细胞的分子机理，并且希望通过与已知结构的NCS2家族蛋白进行比较来探讨此类同向转运蛋白转运底物的结构特征和分子共性，借此加深对同向转运蛋白的了解和认识。

真核生物依赖于在形成糖的器官中能量和碳骨架的有效分配。动物体内的葡萄糖和植物体内的蔗糖是主要的分配形式。细胞内糖的摄取和释放需要囊泡和/或质膜上的转运蛋白。人类和植物用于糖转运的蛋白质来自三个超家族：主要的促进体超家族(MFS)，钠溶解同向转运超家族（SSF；只在动物界）和SWEETs。SWEETs运载单糖和二糖穿过囊泡或质膜。植物的SWEETs在细胞和器官，尤其是在花蜜分泌，长距离转运的韧皮部装载，花粉的营养和种子填充之间的糖转运发挥关键的作用。植物SWEETs可能通过来自感染细胞的糖泄露造成病原体敏感性。有液泡的拟南芥AtSWEET2在根的液泡中隔绝糖。功能缺失的突变体显示增加了对腐霉菌感染的敏感性。.研究中，我们展示了它的同源物，来自水稻( Oryza sativa )的液泡葡萄糖转运体OsSWEET2b，包含一对非对称的三股螺旋束，通过反向连接物跨膜螺旋（TM4）连接以产生转移通路。结构和生化分析显示OsSWEET2b在一个明显的向内（胞质）开放状态形成同源三聚体。TM4与原体的第一个三螺旋束紧密相互作用并介导原体之间的关键连接。来自拟南芥亲近的旁系物SWEET1的结构指导的突变形成识别底物转移和原体串扰的关键残基。了解SWEET1的结构-功能关系对理解真核生物SWEETs的转运机制很有价值，并且可能对工程糖的生产具有一定的指导意义。