Partial purification and characterization of a 60 000-dalton phosphoprotein from pig heart tissue

A 60 000-dalton phosphoprotein (pp60) was purified up to 10(4)-fold by a combination of low-ionic-strength extraction, ammonium sulfate fractionation, on-exchange and affinity chromatography, all in detergent-free buffer. Fractionation on omega-aminohexylagarose column shows that pp60 actually consists of two different polypeptides of similar molecular mass (pp60 omega 1 and pp60 omega 2). Partial hydrolysis with proteases of the proteins 32P-labeled in vitro indicates that pp60 omega 1 and pp60 omega 2 are similar but not identical. On the other hand, individual phosphoamino acid analysis reveals that pp60 omega 1 is phosphorylated primarily at serine residues while pp60 omega 1 is phosphorylated almost equally at serine and threonine residues. Partial hydrolysis with proteases has been also used to explore a possible relationship between the pp60's and the transforming protein of Rous sarcoma virus (pp60v-src). Our data suggest that pp60v-src also consists of two different polypeptides chemically homologous to the presently purified pp60's.