Polymorphism of Platelet Secretory Proteins and Their Mitogenic Activity

Three low molecular weight proteins: β-thromboglobulin (βTG), low affinity plalelet factor 4 (LA-FF4) and platelet basic protein (PBP) have been purified from the material secreted by human platelets stimulated by lonophore A231S7. The preparative steps included heating at 100°C for 5 min, isoelectric focusing (pH gradient 3.5-10.9 in sucrose) and affinity chromatography on hepar in-agarose (elution with 0.4-0.5M NaCl). BTG, LA-PF₄ and PBP showed complete immunological identity using anti-LA-PFi, antibody. Their respective isoelectric points were at pH 7.0, 8.0 and 10.9. Purity was confirmed by NH₂ terminal amino acid analysis (βTG; gly; U1-PF₄ asp; PBP: gly) and electrophoresis. At the NH₂ terminal end LA-PF₄, had four additional amino acids (Asn-Leu-Ala-Lys) while residues 5-21 of LA-PF₄ and 1-16 of βTG were identical. As determined by Isoelectric focusing and specific radioimmunoassay the proportions of βTG, LA-PF₄ and PBP released from platelets were 70%, 14% and 16%, respectively. Evidence is presented that LA-PF₄ is a nrotein originally secreted by human platelets and converted to STG by a heat labile protease associated with the platelets. The three proteins Initiated DMA-synthesis in serum-arrested Swiss 3T3 cells in 0.4% serum. Activity of PBP was observed below 1 ng/ml and saturated at 50 ng/ml. LA-PP₄ and βTG were 10 or 40 fold less active, respectively.