Structural mechanisms for ABC multidrug transporter selectivity and transport

 描述（由申请人提供）：ABC转运蛋白控制药物、脂质和肽在整个人体内的分布，并在多药耐药性中发挥关键作用。该提案旨在了解ABC转运蛋白的整个运输周期，并确定不同ABC转运蛋白如何通过使用X射线晶体学和低温电子显微镜相结合的强大组合方法选择底物的规则。本文概述的方法检查了ABC转运蛋白的机械循环，从底物结合到构象变化，使用冷冻电子显微镜（cryo-EM）方法，该方法在Tomasiak博士与Yifan Cheng博士的合作中产生了ABC转运蛋白前所未有的亚纳米分辨率结构。最终的目标是一个全面的模型，几个人类转运蛋白，揭示了人体药代动力学和药物基因组学的定量模型，这将使这些生物学上的关键分子是如何分布的更清楚的了解。候选人托马斯·托马西亚克（Thomas Tomasiak）有兴趣使用K99开始独立的职业生涯，将多个生物学科和方法结合起来，研究药物、脂质和其他疏水性化合物的分布。Tomasiak博士希望成为领先机构的创新助理教授，为我们在微生物发病机制和癌症中的药物处置和多药耐药性知识做出贡献。该建议以冷冻EM培训计划为中心，该计划将帮助Tomasiak博士成为该领域的专家，并将其与X射线晶体学背景联合收割机相结合。膜蛋白晶体学和cryo-EM的结合将使一个强大的混合方法来回答生物学问题，构象状态的变化和膜蛋白复合物难以单独使用任何技术。拟议的研究的一部分是在开发新的工具，从冷冻EM结构模型的验证。该提案旨在补充托马斯的培训，将他目前的膜蛋白和X射线晶体学知识与新兴的冷冻电子显微镜领域相结合。Tomasiak博士的导师Stroud博士和共同导师Cheng博士在各自的领域都得到了国际认可，Stroud博士支持了许多博士后研究人员向独立过渡。除了共同导师之外，托马西亚克博士还组建了一个由资深科学家组成的咨询委员会，这些科学家将能够帮助指导他的研究，并为他的职业生涯提供理想的指导 发展到独立的地位。

 DESCRIPTION (provided by applicant): ABC transporters govern distribution of drugs, lipids, and peptides throughout the human body and play a critical role in multidrug resistance. This proposal aims to understand the entire transport cycle of an ABC transporter and identify rules governing how different ABC transporters select substrates by using a powerful combination approach combining x-ray crystallography and cryo electron microscopy. The approach outlined here examines the mechanistic cycle of an ABC transporter, from substrate binding to conformational changes using a cryo electron microscopy (cryo-EM) approach that has yielded an unprecedented subnanometer resolution structure of an ABC transporter in a collaboration Dr. Tomasiak was part of with Dr. Yifan Cheng. The ultimate goal is a comprehensive model of several human transporters to reveal a quantitative model of human pharmacokinetics and pharmacogenomics that will give a clearer understanding of how these biologically critical molecules are distributed. The candidate, Thomas Tomasiak, has an interest to use the K99 to launch an independent career weaving several biological disciplines and approaches to studying distribution of drugs, lipids, and other hydrophobic compounds. Dr. Tomasiak hopes to become an innovate assistant professor at a leading institution, contributing to our knowledge of drug disposition and multidrug resistance in microbial pathogenesis and cancer. This proposal is centered on a training program in cryo-EM that will help Dr. Tomasiak become an expert in this field and combine it with his background in x-ray crystallography. The combination of membrane protein crystallography and cryo-EM will make a powerful hybrid approach to answer biological questions concerning conformational state changes and complexes in membrane proteins difficult for either technique alone. Part of the proposed research is in development of new tools for the validation of structural models from cryo-EM. The proposal is designed to supplement Thomas's training by integrating his current knowledge of membrane proteins and x-ray crystallography with the burgeoning field of cryo-electron microscopy. Dr. Tomasiak's mentor, Dr. Stroud, and co-mentor, Dr. Cheng are internationally recognized in their fields and Dr. Stroud has supported numerous postdoctoral researchers into the transition to independence. In addition to the co-mentors, Dr. Tomasiak has assembled an advisory committee of senior scientists who will be able to help guide his research and are ideally situated to guide his career progression to an independent position.