OAT3寡聚状态及编码区非同义单核苷酸多态性对其转运功能影响研究

OAT3是一种药物转运体，在药物的ADME过程中发挥重要作用。然而OAT3可能存在寡聚化并且在中国汉族人群高频发生的编码区非同义单核苷酸多态性，这些蛋白结构差异会影响药物疗效和/或毒性，造成临床药物的组织分布、药代动力学性质改变，产生药物不良反应。前期探索中我们发现OAT3形成了二聚体。本项目将针对OAT3二聚体作用界面及高频SNP位点不清问题，在细胞、蛋白和氨基酸三个层面，研究其蛋白结构与药物转运功能关系。采用蛋白分子印迹、化学交联、荧光能量共转移法判断OAT3二聚体作用界面；采用高表达氨基酸突变的OAT3细胞模型明确OAT3寡聚化氨基酸位点；采用蛋白表达和纯化技术获得高纯度蛋白，通过表面等离子共振技术，明确SNP和临床常用41种药物代谢差异，从而阐明OAT3在个体之间药物反应和毒性差异，为临床用药指导及开发新药物提供理论依据。

OAT3 is a drug transporter which play an important role in the ADME process of drugs. However, there might have been an oligomerization in OAT3, there are many high frequencies single nucleotide polymorphism in Chinese Han nationality. The difference in protein structure can affect the drug efficacy and/or toxicity, thus change the drug tissue distribution and pharmacokinetics, causing the adverse drug reaction. We have found that OAT3 forms a dimer in previous explorations. This study is focusing on the unclear mechanism between oligomerization of OAT3 and high frequency SNP sites and drug metabolism. The relationship between protein structure and drug transport was studied in cell, protein and amino acid level. The oligomerization of OAT3 was determined by western blotting, chemical crosslinking and fluorescence resonance energy transfer. The amino acid sites of OAT3 oligomerization were stated by the cell and model of high expression amino acid mutation OAT3. The high purity protein was obtained by protein expression and purification technology. The metabolic difference between SNP and 41 drugs commonly used in clinic was cleared by surface plasmon resonance. Finally, the individual difference of OAT3 in drug reaction and toxicity was clarified and the theoretical basis for clinical medication guidance and development was provided.