Structural characterization of a human neurotransmitter transporter

人类神经递质转运蛋白的结构表征

• 批准号: 8263767
• 负责人: Kevin H Wang
• 金额: $ 5.39万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-03-03 至 2013-03-02
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8263767
• 关键词: Achievement; Affect; Affinity; Amino Acid Sequence; Amino Acid Transporter; Amino Acids; Amphetamines; Antidepressive Agents; Anxiety; Architecture; Area; Back; Baculoviruses; Binding; Binding Sites; Biogenic Amines; Biological Assay; Brain; Butyric Acids; Carrier Proteins; Cells; Chemical Synapse; Cocaine; Complex; Crystallization; Crystallography; Detergents; Dopamine; Drug Addiction; Embryo; Enzymes; Event; Extracellular Space; Family; Frequencies; Genes; Glycine; Goals; Human; Human Development; Illicit Drugs; Insecta; Lead; Ligands; Lipids; Location; Membrane; Membrane Proteins; Mental disorders; Methods; Molecular; Molecular Structure; Neuroglia; Neurons; Neurotransmitters; Norepinephrine; Orthologous Gene; Pharmaceutical Preparations; Process; Proteins; Psychotropic Drugs; Regulation; Research; Research Training; Resolution; Rewards; Scheme; Scientist; Serotonin; Signal Transduction; Sodium; Specificity; Structural Protein; Structure; Synapses; Therapeutic; Therapeutic Agents; Training; base; clinically relevant; design; gamma-Aminobutyric Acid; inhibitor/antagonist; insight; kidney cell; member; neurotransmitter antagonist; neurotransmitter release; neurotransmitter reuptake; neurotransmitter transport; neurotransmitter uptake; overexpression; presynaptic; protein expression; psychologic; public health relevance; sodium ion; symporter; three dimensional structure; uptake

DESCRIPTION (provided by applicant): The regulation of signaling between neurons depends on the frequency of neurotransmitter release and quantity of neurotransmitter present in the synapse. Efficient termination of signaling is required for resetting the state of the synapse for a subsequent event of neurotransmitter release. The reuptake of neurotransmitter molecules by presynaptic neurons and glial cells depends on membrane-embedded transporter proteins. These enzymes transport neurotransmitters from the extracellular space into the cell and therefore up a concentration gradient. To power this energetically unfavorable process, these transporters also move sodium ions down their concentration gradient and into the cell. Although it is well-established in humans that neurotransmitter transporters directly regulate psychological states such as anxiety and reward-seeking, much less is known about the mechanisms of substrate recognition and inhibition by clinically relevant antagonists. These efforts have been hampered by the absence of atomic-resolution structural information for any eukaryotic neurotransmitter transporters. This application details strategies to express and purify a human neurotransmitter transporter for crystallization and structural determination via x- ray diffraction. The primary goal of this application is to uncover the three-dimensional structure of a neurotransmitter transporter in complex with neurotransmitter and antagonists. This advance will provide the locations of binding sites and generate testable hypotheses to probe how transporters specifically recognize their substrates and are inhibited by antagonists. PUBLIC HEALTH RELEVANCE: Psychoactive drugs and antidepressants are thought to act on neurotransmitter transporters located in neuronal cells in the brain. These transporters normally absorb neurotransmitters back into neuronal cells, and misregulation of this activity can lead to drug addiction and psychological disorders. This application seeks to better understand how drugs affect neurotransmitter transporters by both abused and therapeutic drugs in the hope that more effective compounds can be developed.

描述（由申请人提供）：神经元之间的信号调节取决于神经递质释放的频率和突触中存在的神经递质数量。信号的有效终止是为后续的神经递质释放事件重置突触状态所必需的。突触前神经元和神经胶质细胞对神经递质分子的再摄取依赖于膜嵌入转运蛋白。这些酶将神经递质从细胞外空间转运到细胞内，从而形成浓度梯度。为了给这个能量上不利的过程提供动力，这些转运体也将钠离子沿其浓度梯度移动到细胞内。虽然在人类中，神经递质转运体直接调节心理状态，如焦虑和寻求奖励，但对临床相关拮抗剂对底物识别和抑制的机制知之甚少。由于缺乏任何真核神经递质转运体的原子分辨率结构信息，这些努力受到阻碍。本应用程序详细介绍了通过x射线衍射来表达和纯化人类神经递质转运体的结晶和结构测定的策略。本应用程序的主要目的是揭示神经递质转运体与神经递质和拮抗剂复合物的三维结构。这一进展将提供结合位点的位置，并产生可测试的假设，以探索转运蛋白如何特异性识别其底物并被拮抗剂抑制。