Burr Gary S, Mitchell Cheryl K, Keflemariam Yenabi J, Heidelberger Ruth, O'Brien John
Department of Ophthalmology and Visual Science, University of Texas Health Science Center at Houston, USA.
Biochem Biophys Res Commun. 2005 Oct 7;335(4):1191-8. doi: 10.1016/j.bbrc.2005.08.007.
We examined the interactions of calmodulin with neuronal gap junction proteins connexin35 (Cx35) from perch, its mouse homologue Cx36, and the related perch Cx34.7 using surface plasmon resonance. Calmodulin bound to the C-terminal domains of all three connexins with rapid kinetics in a concentration- and Ca2+-dependent manner. Dissociation was also very rapid. K(d)'s for calmodulin binding at a high-affinity site ranged from 11 to 72 nM, and K(1/2)'s for Ca2+ were between 3 and 5 microM. No binding to the intracellular loops was observed. Binding competition experiments with synthetic peptides mapped the calmodulin binding site to a 10-30 amino acid segment at the beginning of the C-terminal domain of Cx36. The micromolar K(1/2)'s and rapid on and off rates suggest that this interaction may change dynamically in neurons, and may occur transiently when Ca2+ is elevated to a level that would occur in the near vicinity of an activated synapse.
我们利用表面等离子体共振技术研究了钙调蛋白与鲈鱼神经元间隙连接蛋白连接蛋白35（Cx35）、其小鼠同源物Cx36以及相关的鲈鱼Cx34.7之间的相互作用。钙调蛋白以浓度和Ca2+依赖的方式，通过快速动力学与所有三种连接蛋白的C末端结构域结合。解离也非常迅速。在高亲和力位点，钙调蛋白结合的K(d)值范围为11至72 nM，Ca2+的K(1/2)值在3至5 microM之间。未观察到与细胞内环的结合。用合成肽进行的结合竞争实验将钙调蛋白结合位点定位到Cx36 C末端结构域起始处的10 - 30个氨基酸片段。微摩尔级的K(1/2)值以及快速的结合和解离速率表明，这种相互作用在神经元中可能动态变化，并且当Ca2+升高到激活突触附近会出现的水平时可能会短暂发生。
