Itakura M, Iio T
Department of Physics, Faculty of Science, Nagoya University, Aichi.
J Biochem. 1992 Aug;112(2):183-91. doi: 10.1093/oxfordjournals.jbchem.a123875.
Ca2+ binding to calmodulin triggers conformational change of the protein which induces exposure of hydrophobic surfaces. Melittin has been believed to bind to Ca(2+)-bound calmodulin through the exposed hydrophobic surfaces. However, tryptophan fluorescence measurements and gel chromatography experiments with the melittin-calmodulin system revealed that melittin bound to calmodulin at zero salt concentration even in the absence of Ca2+; addition of salt removed melittin from Ca(2+)-free calmodulin. This means not only the hydrophobic interaction but also the electrostatic interaction contributes to the melittin-calmodulin binding. The fluorescence stopped-flow studies of the dissociation reaction of melittin-calmodulin complex revealed that Ca2+ removal from the complex induced a conformational change of calmodulin, resulting in reduction of the hydrophobic interaction between melittin and calmodulin, but the electrostatic interaction kept melittin still bound to calmodulin for a subsecond lag period, after which melittin dissociated from calmodulin. The fluorescence stopped-flow experiments on the dissociation reaction of complex of melittin and tryptic fragment(s) of calmodulin revealed that the lag period of the melittin dissociation reaction was attributable to the interaction between the C-terminal half of calmodulin and the C-terminal region of melittin.
钙离子与钙调蛋白结合会引发该蛋白的构象变化，进而导致疏水表面暴露。人们一直认为蜂毒肽通过暴露的疏水表面与结合了钙离子的钙调蛋白结合。然而，对蜂毒肽 - 钙调蛋白系统进行的色氨酸荧光测量和凝胶色谱实验表明，即使在没有钙离子的情况下，蜂毒肽在零盐浓度下也能与钙调蛋白结合；添加盐会使蜂毒肽从无钙离子的钙调蛋白上脱离。这意味着不仅疏水相互作用，静电相互作用也对蜂毒肽 - 钙调蛋白的结合有贡献。对蜂毒肽 - 钙调蛋白复合物解离反应的荧光停流研究表明，从复合物中去除钙离子会引发钙调蛋白的构象变化，导致蜂毒肽与钙调蛋白之间的疏水相互作用减弱，但静电相互作用使蜂毒肽在亚秒级的延迟期内仍与钙调蛋白结合，之后蜂毒肽才从钙调蛋白上解离。对蜂毒肽与钙调蛋白胰蛋白酶片段复合物解离反应的荧光停流实验表明，蜂毒肽解离反应的延迟期归因于钙调蛋白的C端半段与蜂毒肽的C端区域之间的相互作用。
