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NBCe1 and n1: Crystal Structure Determination

NBCe1 和 n1：晶体结构测定

基本信息

批准号：
7111304
负责人：
Harry Singh Gill
金额：
$ 5.4万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-09-01 至 2008-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7111304
关键词：
Saccharomyces cerevisiae X ray crystallography Xenopus acidity /alkalinity bicarbonates crystallization electrophysiology electroporation gene expression ion transport membrane transport proteins method development postdoctoral investigator protein purification protein structure function sodium

项目摘要

DESCRIPTION (provided by applicant): Bicarbonate (HCO3-) transport across epithelial cells of the kidney is of great importance to maintain systemic acid-base balance. The HCO3- transporters NBCe1 & NBCn1 are integral, membrane-proteins that, respectively, play important roles in HCO3- reabsorption in the proximal tubule and NH4+ reabsorption in the thick ascending limb. The specific aims of this project are: (1) To use X-ray crystallography to illuminate the structures of the N-terminal domains of NBCe1 and n1; (2) To explore the physiological roles in NBCn1 of the unique 124-residue exon 7, which may be spliced in or out of the protein; (3) To develop a large-scale, eukaryotic expression system for the membrane domain of NBCe1 or NBCn1, to be used later for crystallization trials. My preliminary work on the purification and crystallization of the N-terminus of NBCe1 is paving the way for vital structural information needed to study the regulation of HCO3- transport, and my new splice variants of NBCn1 broadens our current understanding of the role of NBCn1 in various cell types. The proposed study will provide key insights on the molecular mechanisms of renal HCO3- transport and will enhance our understanding of processes fundamental to renal disease.

描述（由申请方提供）：跨肾脏上皮细胞的碳酸氢盐（HCO 3-）转运对于维持全身酸碱平衡非常重要。HCO 3转运蛋白NBCe 1和NBCn 1是不可或缺的膜蛋白，分别在近端小管中的HCO 3重吸收和粗升支中的NH 4+重吸收中发挥重要作用。本项目的具体目标是：（1）利用X射线晶体学方法研究NBCe 1和n1的N端结构域：（2）探讨NBCn 1中独特的124个氨基酸的外显子7的生理功能，该外显子7可能被剪接进或剪接出蛋白质;（3）开发NBCe 1或NBCn 1膜结构域的大规模真核表达系统，用于以后的结晶试验。我对NBCe 1 N-末端的纯化和结晶的初步工作为研究HCO 3-转运调控所需的重要结构信息铺平了道路，我的NBCn 1新剪接变体拓宽了我们目前对NBCn 1在各种细胞类型中作用的理解。这项研究将为肾脏HCO 3转运的分子机制提供关键的见解，并将增强我们对肾脏疾病基本过程的理解。