Ion-selectivity and Mechanisms of P-type IIC ATPases

智力优势所有细胞都能维持跨质膜的离子梯度，为多种生理过程提供能量。这种梯度由Na/K泵（一种P型2 ATP酶）建立和维持。这类蛋白质的成员存在于所有门和运输不同的金属几乎相同的机制。离子识别和离子驱动的构象变化是这种通用机制的重要组成部分，这种机制赋予每个ATP酶独特性，但人们对此知之甚少。该项目的重点是P型IIC ATP酶，所有动物表达的蛋白质，其功能对于适应不同的盐度，消化和细胞内稳态至关重要。该研究将使用分子生物学，高时间分辨率电生理学和荧光技术相结合的方法来研究允许P型IIC ATP酶识别运输离子的分子机制以及离子运输如何受蛋白质构象变化控制。特别是，来自P型IIC ATP酶的晶体结构的结构信息将用于靶向结合离子的区域和蛋白质的细胞内区域中的变化，通过电压钳技术结合荧光监测功能影响。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。更广泛的影响这个项目的更广泛的影响目标是激励本科生追求基础基础科学的职业生涯，并通过科学研究发展技术技能和对发现的态度。PI的实验室有着让本科生和研究生参与科学发现的历史，受训人员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。一些受训人员从事科学工作。该项目将涉及来自不同课程和学科的本科生，包括重点关注从平原桥梁到学士学位课程等机构课程中招募代表性不足的少数民族。该项目的成果将通过在影响力大的期刊上发表文章、举办课程、应邀在知名度高的国家和国际科学会议上发表演讲和介绍等方式传播。

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.