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Abstract: Asp49 plays a key role coordinating calcium in the catalytic site of phospholipases A2 (PLA2’s), thus assisting the stabilization of the ES intermediary transition state, which induces the enzyme activation. In this work, molecular dynamics simulations of an acidic PLA2 from the venom of Agkistrodon halys pallas were performed in water, methanol and octanol. Because of the low affinity of svPLA2’s for calcium ion in aqueous systems, different positional restraint forces were applied and the coordinates describing svPLA2–Ca2+ ligand properties were simulated and evaluated. The results of the simulations were used to propose an interfacial activation model for svPLA2’s. In this model, events related with enzyme activation involve: (a) the reorganization of calcium binding loop at membrane proximity followed by the Ca2+ uptake; (b) side chain reorientation of Trp31, which defines a new specificity pocket for the phospholipid chain; (c) reduction of the distance between His48 and Asp49, increasing the nucleophilicity of Nε−His48; (d) side chain reorientation of Lys69 concomitant with projection of the 69-loop to solvent. In addition to existing biochemical and crystallographic data, our results allow us to describe a more detailed model for the interfacial activation of phospholipases A2. [Copyright &y& Elsevier]