蛋白酪氨酸磷酸酶非受体型12与心脏HERG钾通道相互作用并调控通道功能的研究-内科学专业论文.docxVIP

汕头 汕头大学医学院硕士研究生毕业论文 II II 结果：（1）免疫共沉淀分析发现抗 HERG 的抗体能够沉淀 HERG 和 PTPN12 复合物；（2） 应用 GST pull-down 分析发现 GST-HERG-NT 能够将 PTPN12 沉淀，而 GST 蛋白则不能沉 淀 PTPN12；（3）免疫荧光组织化学分析：用倒置荧光显微镜观察可见在细胞膜上 PTPN12 蛋白和 HERG 蛋白分别发出绿色荧光和红色荧光，二者荧光重叠部分（黄色荧光）提示 这两种蛋白在细胞上共定位，发现 PTPN12 和 HERG 两个蛋白共定位的地方主要出现在 细胞膜。（4）膜片钳检测发现，PTPN12 蛋白能降低 HERG 电流幅度。 结论：本实验结果表明蛋白酪氨酸磷酸酶非受体型 12（PTPN12）与心脏 HERG 钾通道 的氨基末端存在相互作用，并通过这种相互作用发挥了调控 HERG 钾通道功能的作用， 使 HERG 钾通道电流振幅降低。本结果提示 PTPN12 参与了 HERG 钾通道功能的调控， 其机制可能是通过改变 HERG 钾通道磷酸化程度而发挥作用。这一发现将有助于进一步 理解 HERG 通道功能的调控机制，为更深刻理解 LQTS 的发病机理及开发治疗 LQTS 的 蛋白质药物提供新的思路。 关键词：HERG 钾通道、蛋白酪氨酸磷酸酶非受体型 12、免疫共沉淀、GST pull-down Assay、 荧光免疫组织化学；膜片钳技术 III III Abstract Background: Encoded by the Human ether-a-go-go-related gene (HERG), HERG channel belongs to the voltage-gated channels and mediates the rapid component of the cardiac delayed rectifier current (IKr). HERG channel underlies the regular cardiac electric activity. Mutations in the HERG gene and the blockade of the HERG-coded potassium channel by drugs cause the long QT syndrome. HERG channel underlies the regular cardiac electric activity. Mutations in the HERG gene and the blockade of the HERG-coded potassium channel by drugs cause the long QT syndrome. In recent years, ten genes have been reported to correlate with LQTS. The mutations in HERG gene channel cause type 2 long QT syndrome(LQT2)，which are the second common reason of LQTS. Protein - protein interaction is an important basis for many cellular functions, for exemple ,signal transduction, cell cycle regulation, RNA transcription, DNA replication, protein translation, protein post-translational processing and modification functions are dependent on the completion of protein - protein interactions.in recent years ,several cellular proteins have been reported to interact with HERG channel and modify its expression, trafficking and function. Fifteen proteins ， including PTPN12 （ Protein Tyrosine-protein phosphatase non-receptor type 12 ）， caveolin-1;FHL2;Myotrophin,