C2A activates a cryptic Ca 2+ -triggered membrane penetration activity within the C2B domain of synaptotagmin I (original) (raw)

2002, Proceedings of the National Academy of Sciences

Synaptotagmin (syt) I, an integral membrane protein localized to secretory vesicles, is a putative Ca 2+ sensor for exocytosis. Its N terminus spans the membrane once, and its cytoplasmic domain contains two conserved C2 domains, designated C2A and C2B. The isolated C2A domain penetrates membranes in response to Ca 2+ ; isolated C2B does not. Here, we have addressed the function of each C2 domain, but in the context of the intact cytoplasmic domain (C2A-C2B), by using fluorescent reporters placed in the Ca 2+ -binding loops of either C2A or C2B. Surprisingly, these reporters revealed that, analogous to C2A, a Ca 2+ -binding loop in C2B directly penetrates into lipid bilayers. Penetration of each C2 domain was very rapid ( k on ≈10 10 M −1 ⋅s −1 ) and resulted in high affinity C2A-C2B–liposome complexes ( K d ≈13–14 nM). C2B-bilayer penetration strictly depended on the presence, but not the membrane binding activity, of an adjacent C2A domain, severing C2A from C2B after protein synt...

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