Ion-selectivity and Mechanisms of P-type IIC ATPases

所有细胞都在质膜上维持离子梯度，为多种生理过程提供能量。这种梯度是由Na/K泵（一种p型2 atp酶）建立和维持的。该蛋白类的成员存在于所有门中，并以几乎相同的机制运输不同的金属。离子识别和离子驱动的构象变化是这种赋予每个atp酶奇点的普遍机制的重要组成部分，但人们对这种机制知之甚少。本项目重点研究p型IIC atp酶，这是一种所有动物都表达的蛋白质，其功能对适应不同盐度、消化和细胞稳态至关重要。该研究将采用分子生物学、高时间分辨率电生理学和荧光技术相结合的方法，研究p型IIC atp酶识别转运离子的分子机制，以及蛋白质构象变化如何控制离子转运。特别是，来自p型IIC atp酶晶体结构的结构信息将用于靶向结合离子区域和蛋白质胞内区域的变化，并通过结合荧光的电压钳技术监测功能影响。该项目的结果将提供关键的见解，并有望解决有关p型atp酶超家族分子力学的流行模型。更广泛的影响本项目更广泛的影响目标是激励本科生追求基础科学的职业生涯，并通过科学研究培养技术技能和对发现的态度。PI的实验室有让本科生和研究生参与科学发现的历史，受研者在高影响力期刊上担任文章的共同作者，并在国家科学会议上展示他们的发现。一些受训者从事科学事业。该项目将涉及来自不同专业和学科的本科生，包括从机构项目（如平原桥梁到学士学位项目）中招募代表性不足的少数民族。该项目的成果将通过在高影响力期刊、课程、特邀讲座和在高知名度的国家和国际科学会议上的演讲等出版物进行传播。

INTELLECTUAL MERITAll cells maintain ion gradients across the plasma membrane, which provides energy for multiple physiological processes. This gradient is established and maintained by the Na/K pump, a P-type 2 ATPase. Members of this protein class are present in all phyla and transport different metals with nearly identical mechanisms. Ion recognition and ion driven conformational changes are an essential part of this universal mechanism that gives singularity to each ATPase and are poorly understood. This project focuses on P-type IIC ATPases, proteins expressed by all animals, whose function is essential for adaptation to different salinities, digestion and cellular homeostasis. The research will use a combined approach of molecular biology, high-temporal resolution electrophysiology and fluorescence techniques to study the molecular mechanisms that allow recognition of the transported ions by P-type IIC ATPases and how ion transport is controlled by protein conformational changes. In particular, structural information from crystal structures of P-type IIC ATPases will be used to target changes in the regions that bind ions and in intracellular regions of the proteins, with functional affects monitored by voltage clamp techniques combined with fluorescence. The results from this project will provide critical insight and hopefully resolve prevailing models regarding the molecular mechanics of the P-type ATPase superfamily. BROADER IMPACTSThe broader impacts goal of this project is to inspire undergraduate students to pursue careers in fundamental basic sciences, and to develop the technical skills and attitudes towards discovery through scientific research. The PI's laboratory has a history of involving undergraduate and graduate students in scientific discovery, with trainees serving as co-authors on articles in high-impact journals, as well as presenting their findings at national scientific meetings. Several of trainees have pursued scientific careers. This project will involve undergraduate students from various programs and disciplines, including a focus on the recruitment of underrepresented minorities from institutional programs such as the Plains Bridges to Baccalaureate program. The results from this project will be disseminated by publications in high-impact journals, courses, invited lectures and presentations at high visibility national and international scientific meetings.