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The location of the membrane-docking face on the Ca2+-activated C2 domain of cytosolic phospholipase A was investigated. The experiment utilized 16 engineered cysteines that were individually substituted for solvent-exposed residues spanning all surface regions of the cPLA2 C2 domain. Results showed that docking of C2 domains is stimulated by the binding of multiple Ca2+ ions to a conserved set of residues found within three otherwise variable Ca2+-binding loops.

The role of cytosolic phospholipase A2 (cPLA2 C2) domain in the Ca2+ ion binding was determined through utilization equilibrium binding and stopped-flow kinetic experiments. It was shown that Ca2+-regulated C2 domains typically bind two Ca2+ ions with positive cooperativity resulting in conformational change and membrane docking. Intrinsic fluorescence of Trp (super 71) was enhanced following binding of Ca2+ to the domain in the