Characterization of human platelet basic protein, a precursor form of low-affinity platelet factor 4 and .beta.-thromboglobulin

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Abstract
Platelet basic protein (PBP) was purified from the supernatant of thrombin-stimulated, washed human platelets by ion-exchange, affinity, molecular sieve, and high-performance liquid chromatography (HPLC). The NH2-terminal amino acid sequence was determined by automated Edman degradation, revealing 9 unique residues followed by 10 residues of the established low-affinity platelet factor 4/.beta.-thromboglobulin (LA-PF4/.beta.TG) sequence. Among the nine were three basic residues, accounting for the high isoelectric point of PBP. Additional evidence for precursor status includes the immunological cross-reactivity of all three species and the ability of plasmin and trypsin to produce from PBP a species resembling .beta.TG in charge, hydrophobicity, and size. Tryptic peptide maps of PBP and LA-PF4 obtained by reverse-phase HPLC were very similar, and from each protein, a peptide was isolated which showed the amino acid composition predicted for the COOH-terminal tryptic peptide of .beta.TG. Normal platelets contained predominantly LA-PF4, with PBP ranging from 10% to 30% of total .beta.TG antigen. This was true even when fresh platelets were lysed with trichloroacetic acid in order to provide the most complete and rapid inhibition of proteolytic activity. .beta.TG itself was never detected in this situation or in the release supernatant of stimulated platelets, and only rarely in unprotected lysates. In agreement with earlier results, crude preparations of PBP were mitogenic for 3T3 cells, but highly purified preparations of PBP and LA-PF4 were free of this activity.