Fig. 3.

Apparent Ca²⁺ affinities of wild-type and mutant single and double C₂ domain fragments from synaptotagmin 1 measured by Ca²⁺-dependent binding to liposomes. The isolated wild-type and D232N and D238N mutant C₂A domain, the wild-type C₂B domain, and the wild-type and mutant double C₂ domain fragments were analyzed. Liposomes composed of 25% PS/75% PC were incubated with the indicated C₂ domains (present as soluble purified GST-fusion proteins) at the Ca²⁺ concentrations shown on top (clamped with Ca²⁺/EGTA buffers) and centrifuged, and bound proteins were analyzed by SDS-PAGE and Coomassie blue staining. Data shown are from a single representative experiment repeated multiple times.