Purification and characterization of calpastatin from grass prawn muscle (Penaeus monodon)

Calpastatin, a specific calpain inhibitor was purified to electrophoretical homogeneity from grass prawn (Penaeus monodon) muscle by 100 degrees C hea t-treatment, DEAE-Sephacel, and Q-Sepharose chromatographs. No significant change in the inhibitory activity of crude calpastatin was observed even af ter 20 min incubation at 100 degrees C, pH 7.0. The purified prawn calpasta tin had a molecular weight (M-r) of 80 and 88.7 kDa determined by SDS-PAGE and Sephacryl S-200 HR gel filtration, respectively. According to the activ e site titration, the purified calpastatin revealed four beef mu-calpain an d two beef m-calpain binding domains, respectively. It was stable during 1 h of incubation at 30 degrees C under pH 4.5-10.0 and shown to be a highly specific inhibitor for calpain.