Davletov B A, Südhof T C
Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas 75235.
J Biol Chem. 1994 Nov 18;269(46):28547-50.
Synaptotagmin I is a Ca2+/phospholipid binding protein of synaptic vesicles with a proposed function as a Ca2+ sensor in synaptic vesicle exocytosis. Using controlled partial proteolysis as an assay, we now show that synaptotagmin I undergoes a conformational change as a function of Ca2+ binding. As observed for phospholipid binding, Ba2+ and Sr2+ but not Mg2+, substitute for Ca2+ in effecting this conformational change. The first C2 domain from synaptotagmin I that represents the Ca(2+)-dependent phospholipid binding domain of synaptotagmin also undergoes a Ca(2+)-dependent change in controlled partial proteolysis. In contrast, no effect of Ca2+ was observed with mutant C2 domains containing point mutations that abolish Ca2+ binding. The Ca2+ concentration dependence of the effect of Ca2+ on proteolysis mirrors the Ca2+ dependence of phospholipid binding. The conformational shift in synaptotagmin I caused by Ca2+/phospholipid binding could be the basis for its Ca(2+)-regulated function in triggering neurotransmitter release.
突触结合蛋白I是一种突触小泡的Ca2+/磷脂结合蛋白，在突触小泡胞吐作用中被认为具有Ca2+传感器的功能。我们现在利用可控的部分蛋白酶解作为一种检测方法，证明突触结合蛋白I会随着Ca2+结合而发生构象变化。正如在磷脂结合中观察到的那样，Ba2+和Sr2+而非Mg2+，在引起这种构象变化时可替代Ca2+。突触结合蛋白I的第一个C2结构域代表突触结合蛋白的Ca(2+)依赖性磷脂结合结构域，在可控的部分蛋白酶解中也会发生Ca(2+)依赖性变化。相比之下，对于含有消除Ca2+结合的点突变的突变C2结构域，未观察到Ca2+的影响。Ca2+对蛋白酶解作用的浓度依赖性反映了磷脂结合的Ca2+依赖性。由Ca2+/磷脂结合引起的突触结合蛋白I的构象转变可能是其在触发神经递质释放中Ca(2+)调节功能的基础。
