Calcium-dependent proteolysis of actin during storage of platelet concentrates

In an ongoing study of the changes that occur in platelet concentrates during storage, we investigated two 28–26-Kd proteins designated SP–1 and SP–2, respectively, which increase markedly during blood-bank storage of platelet concentrates at room temperature. Formation of SP–1 and SP–2 was inhibited by storage at 4 degrees C as well as by treatment of the concentrates with leupeptin, N–ethylmaleimide, and EDTA; DFP and PPACK had no effect. The calcium ionophore A23187 markedly stimulated production of SP–1 and SP–2. After partial purification, the two proteins were found to be associated with platelet cytoskeletal protein. Two-dimensional peptide mapping and amino acid sequencing identified SP-1 and SP-2 as fragments of actin formed by cleavage on the N-terminal side of residues Thr–106 and Ala- 114, respectively. Generation of SP–1 and SP–2 during storage of platelet concentrate is likely attributable to calcium-dependent neutral protease degradation of actin and may have implications for development of the platelet storage lesion.