Structure and Function of Concentrative Nucleoside Transporters

浓缩核苷转运蛋白的结构和功能

• 批准号: 9024572
• 负责人: Seok-Yong Lee
• 金额: $ 29.41万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-05-01 至 2017-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9024572
• 关键词: Adenosine; Adopted; Adverse effects; Amino Acids; Antineoplastic Agents; Antiviral Agents; Ara-C; Architecture; Binding; Binding Sites; Biological Assay; Biological Availability; Biological Models; Cancer cell line; Cell membrane; Cells; Chemicals; Complex; Coupled; Coupling; DNA; Data; Detergents; Development; Drug Targeting; Drug Transport; Electrodes; Family; Goals; Health; Homologous Gene; Human; Human body; Integral Membrane Protein; Intestines; Ions; Kidney; Lighting; Liver; Membrane; Molecular; Molecular Conformation; Mutation; Nucleic Acids; Nucleoside Transporter; Nucleosides; Organ; Organism; Pharmaceutical Preparations; Pharmacology; Pharmacotherapy; Physiological; Physiology; Positioning Attribute; Property; Protein Engineering; RNA chemical synthesis; Radioactive; Reagent; Resistance; Ribavirin; Role; Sequence Homology; Signal Transduction; Sodium; Source; Specificity; Structure; Substrate Specificity; Testing; Uridine; Vibrio cholerae; Zidovudine; base; cancer cell; cancer type; cytotoxic; design; gemcitabine; improved; insight; mutant; novel therapeutics; protein transport; research study; response; screening; solute; stoichiometry; voltage clamp

DESCRIPTION (provided by applicant): Nucleoside transporters (NTs) are integral membrane proteins that are designed to transport nucleosides into the human body. Nucleoside transport across cell membranes is physiologically important as it provides the major source of nucleosides for DNA/RNA synthesis in cells and it is responsible for the termination of adenosine signaling. Another important fact about NTs is that many nucleoside-derived anticancer and antiviral drugs are transported into cells by way of NTs. Despite their importance in physiology and pharmacology, the molecular mechanism of NTs is largely unknown due to the lack of atomic structures of the transporters. Great advances in the field depend on the structure determination and illumination of the transport mechanism of NTs. Our long- term goal is to understand the fundamental mechanism of selective nucleoside transport by concentrative nucleoside transporters (CNTs) with structural and functional studies. CNTs belong to a family of the solute carrier transporter superfamily (SLC28) and use an ion gradient to transport nucleosides actively against their chemical gradients. We have chosen to use a CNT homolog from Vibrio cholerae (vcCNT) as a model system for our structural and functional studies. vcCNT is an excellent model system to study hCNTs that shares significant sequence homology and functional properties with hCNTs. We solved the crystal structure of vcCNT in complex with uridine. The structure reveals the overall architecture and the nucleoside-binding site of the transporter, providing mechanistic insights of selective nucleoside transport by vcCNT and CNT in general. These new results have led us to propose a combined structure-function study with the following three aims: 1) Determination of the crystal structures of vcCNT in complex with different nucleosides and nucleoside drugs, 2) Functional characterization and comparison of vcCNT with hCNTs, and 3) Structure determination of vcCNT in alternate conformations. The proposed studies will significantly advance our understanding of the mechanism of nucleoside and nucleoside drug transport by prokaryotic and eukaryotic CNTs, which eventually will facilitate the development of new therapeutic reagents.

描述（由申请人提供）：核苷转运蛋白（NTS）是旨在将核苷转运到人体的整体膜蛋白。核苷跨细胞膜的转运在生理上很重要，因为它为细胞中DNA/RNA合成的核苷提供了主要来源，并且它负责终止腺苷信号传导。关于NTS的另一个重要事实是，许多核苷衍生的抗癌药和抗病毒药物通过NTS转运到细胞中。尽管它们在生理学和药理学中的重要性，但由于缺乏转运蛋白的原子结构，NTS的分子机制在很大程度上是未知的。该领域的巨大进展取决于NTS运输机制的结构确定和照明。我们的长期目标是通过结构和功能研究来了解浓缩核苷转运蛋白（CNT）选择性核苷转运的基本机制。 CNT属于溶质载体转运蛋白超家族（SLC28）的家族，并使用离子梯度对其化学梯度进行积极运输核苷。我们选择使用来自Vibrio Cholerae（VCCNT）的CNT同源物作为我们结构和功能研究的模型系统。 VCCNT是研究HCNT的出色模型系统，该系统具有与HCNT相同的序列同源性和功能性能的重要模型系统。我们在与尿苷中求解了VCCNT的晶体结构。该结构揭示了转运蛋白的整体结构和核苷结合位点，从而提供了通过VCCNT和CNT的选择性核苷转运的机械见解。这些新结果使我们提出了一项结合结构功能研究，其目的是：1）确定与不同核苷和核苷药物复合物中VCCNT的晶体结构，2）2）VCCNT与HCNT的功能表征和比较，以及3）交替构象中VCCNT的结构确定。拟议的研究将显着提高我们对原核生物和真核CNT核苷和核苷药物转运机制的理解，这最终将促进新的治疗试剂的发展。