LIGAND BINDING SITES ON THE DOPAMINE TRANSPORTER

描述：（申请人摘要） 开发治疗可卡因滥用的药物是一个重要目标。 在 为了开发阻止可卡因与其靶点结合的新药， 多巴胺转运蛋白（DAT），这将是非常有用的，知道在哪里， 可卡因和多巴胺结合在转运蛋白上。 的一种方法 可卡因结合位点的定位一直是为了探索 影响与转运蛋白结合的结构特征。 延长 这种方法的一个重要方面是开发出一种抑制剂， 传送器 这类化合物与蛋白质反应形成 抑制剂和蛋白质之间的共价键。 因此，目标是 来鉴定共价连接的位点。 这是通过分裂 蛋白质和随后的蛋白质片段，配体保留在其上 附 NIDA的研究人员已经使用这种方法开始 定位两个光亲和配体，[125 I]RTI-82，可卡因类似物，和 [125 I]DEEP，一种DAT不同结构域的GBR类似物。 拟议的工作旨在使用质谱法来识别这些位点 这些配体在先前确定的DAT中的附着 片段 此外，两个新的光亲和性的附着位点 也将鉴定作为DAT底物的配体。 的 综合结果将有助于构建配体结合的图像 DAT功能和抑制的关键区域。

DESCRIPTION: (Applicant's Abstract) Development of medications to treat cocaine abuse is an important goal. In order to develop new drugs which prevent binding of cocaine to its target, the dopamine transporter (DAT), it would be very useful to know where both cocaine and dopamine bind on the transporter protein. One approach to localization of the binding site for cocaine has been to explore the structural features which affect binding to the transporter. An extension of this approach has been to develop inhibitors which bind irreversibly to the transporter. This class of compounds reacts with the protein to form a covalent bond between the inhibitor and the protein. The objective then is to identify the site of covalent attachment. This is done by cleavage of the protein and following the protein fragment to which the ligand remains attached. Work by investigators at NIDA have used this method to begin to localize two photoaffinity ligands, [125I]RTI-82, a cocaine analog, and [125I]DEEP, a GBR analog to different domains of DAT. The proposed work is designed to use mass spectrometry to identify the sites of attachment of these ligands within the previously identified DAT fragments. In addition, the sites of attachment of two new photoaffinity ligands, which are substrates for DAT, will also be identified. The combined results will help to construct a picture of the ligand binding regions critical for DAT function and inhibition.