Purification and preliminary characterization of a plasma kallikrein inhibitor isolated from sea hares Aplysia dactylomela Rang, 1828

An inhibitor active against pancreatic trypsin was found in the crude extract from the sea hares Aplysia dactylomelaRang, 1828. A stronger inhibitory activity against human plasma kallikrein was detectable after treating this extract at 60 °C, for 30 min. The plasma kallikrein inhibitor (AdKI) purification was achieved by acetone fractionation (80%) v/v, ion-exchange chromatography on Mono Q column and gel filtration chromatography on Superdex 75 column (FPLC system). By the latter a molecular mass of 2,900 Da was estimated. The purified inhibitor strongly inhibits human plasma kallikrein with a K i value of 2.2×10 −10 M, while human plasmin and pancreatic trypsin were inhibited with K i values of 1.8×10 −9 and 4.7×10 −9 M, respectively. Chymotrypsin, pancreatic elastase, pancreatic kallikrein and thrombin are not inhibited. The effect of AdKI on plasma kallikrein was confirmed by the prolongation of activated partial thromboplastin time, using a clotting time assay. The inhibitor did not affect prothrombin time or thrombin time. AdKi is a more specific inhibitor than other serine proteinase inhibitors from marine invertebrates.