PICK1中N端酸性区域对其PDZ结构域脂质结合能力的调节

蛋白激酶Cα相互作用蛋白1(protein interacting with Cα kinase 1, PICK1)是衔接膜上受体和蛋白激酶Cα的重要蛋白.利用荧光光谱结合定点突变技术 、蛋白与脂质覆盖法等方法，分析了PICK1蛋白N末端区域几个酸性氨基酸残基对PDZ 结构域与膜脂结合的影响，以及钙离子结合N末端酸性区域对PDZ脂结合能力的调节. 结果显示, 带有上游酸性区域的PDZ结构域（NPDZ）的脂质结合能力仅相当PDZ结构 域的15%，相比单独的PDZ结构域与脂质的解离常数Kd(PDZ)为1.58×103 μg·L-1， NPDZ与脂质解离常数Kd(NPDZ)为3.3×104μg·L-1，其中在N末端酸性残基中D8与 D12两个天冬氨酸是影响脂质结合能力减弱的关键残基，若将二者分别突变为丙氨酸 后,NPDZ与脂质的解离常数分别为：Kd (D8/A)=4.42×103μg·L-1；Kd (D12/A) =1.73×103μg·L-1接近于PDZ结构域与脂质结合能力；钙离子会增强NPDZ脂结合能力，当钙离子浓度达到30 μmol/L时，NPDZ的脂结合能力提高2.3倍，但只相当于PDZ的50% 的结合能力.

Regulation of Liposome Binding of PDZ Domain by Its Adjacent N-terminal Acidic Region

Protein interacting with Cα kinase 1 (PICK1), plays an important role in linking the membrane receptors and PKCα in synapse. We have analyzed the interaction between the PDZ domain and membrane lipid regulated by its adjacent N-terminal acid region, using fluorescence spectra, gene manipulation combined with PLO (protein-lipid overlay assay). The result indicated that N-terminal acid region of PICK1 decreased the interaction between PDZ domain of PICK1 and membrane lipid about 85% degree. D8, D12 of N-terminal acidic regions constitute the key residues for decreasing PDZ binding to lipid. 30 μmol/L Ca2+ could enhance the lipid binding ability of PDZ domain with its adjacent N- terminal acid region about 2.3 times, but that was about 50% degree compared to that of only PDZ domain.