Structural Biology of the Apical Bile Acid Transporter

胆汁酸被认为是第二信使和荷尔蒙，与 血糖、脂肪和能量代谢的临床意义。人体顶端的钠- 依赖胆汁酸转运体(HASBT)在肠-肝循环中起关键作用 胆盐，胆固醇动态平衡，并作为分子靶点 高胆固醇药物和药物前药策略。尽管它在临床上 重要的是，ASBT在分子水平上的特征很差。建议数 研究将集中在ASBT的结构生物学上。两个最新的细菌晶体 结构声称与hASBT有进化关系，但它们的拓扑结构形成了鲜明的对比 到文献中的生化数据。当前的提案直接面对这一点 争议和我们推进胆汁酸转运体结构理解的计划， 功能和调节是围绕以下目标组织的：1)做细菌 转运蛋白结构为真核生物提供了具有代表性的结构模型 SLC10同源基因？我们假设对基因祖先的分析和 同源基因的底物表型可以决定进化和结构的关联性 (假定)原核和真核SLC10A家族成员；此外，我们的目标是 真核细胞SLC10A的表达、纯化、结晶及X-射线结构测定 同源同源基因；2)描述和绘制其功能调控和调控图谱 HASBT表达；我们将应用蛋白质组学方法来确定后 时间上的翻译修饰。根据我们的初步数据，我们将进一步 研究高阶二聚体或多聚体中hASBT的组织作为一种方法 控制功能。通过这些研究获得的信息将显著增加我们的 对驱动胆汁酸转运的结构相互作用的理解 一般的溶质载体(SLC)蛋白质的结构知识。此外，它可能有助于 抗高胆固醇血症的特效治疗策略的未来发展 相关的心血管和代谢性疾病。

Bile acids have been recognized to function as second messengers and hormones, with clinical implications in glucose, lipid and energy metabolism. The human apical sodium- dependent bile acid transporter (hASBT) plays a key role in the enterohepatic recycling of bile salts, cholesterol homeostasis, and serves as a molecular target for hypercholesterolemic agents and pharmaceutical prodrug strategies. Despite its clinical significance, ASBT is poorly characterized at the molecular level. The proposed research will focus on the structural biology of ASBT. Two recent bacterial crystal structures claim evolutionary relatedness to hASBT, yet their topology is in clear contrast to biochemical data in the literature. The current proposal directly confronts this controversy and our plan to advance understanding of bile acid transporter structure, function and regulation is organized around the following aims: 1) Do bacterial transporter structures provide representative structural models for eukaryotic SLC10 orthologs? wherein we hypothesize that analysis of genetic ancestry and substrate phenotype of orthologs can determine evolutionary and structural relatedness of (putative) prokaryotic and eukaryotic SLC10A family members; further, we aim to express, purify, crystallize and determine the x-ray structure of eukaryotic SLC10A orthologs; 2) To characterize and map the functional regulation and modulation of hASBT expression; where we will apply a proteomics approach to determining post- translational modifications temporally. Based on our preliminary data we will further investigate the organization of hASBT in higher-order dimers or multimers as a way of controlling function. Information gained by these studies will significantly increase our understanding of the structural interactions that drive bile acid transport and further our structural knowledge of solute carrier (SLC) proteins in general. Additionally, it may aid future development of specific therapeutic strategies against hypercholesterolemia and related cardiovascular and metabolic diseases.