Physical modification of purified Rous sarcoma virus pp60v-src protein after incubation with ATP/Mg2+.

Using partially purified enzyme preparations, we show that incubation of the Rous sarcoma virus transforming protein kinase, pp60v-src, with Mg2+ and ATP at concentrations near or above the enzyme's Km for ATP resulted in a physical modification of the pp60v-src polypeptide. Under such conditions, a portion of pp60v-src was converted to a form that migrated more slowly in SDS-polyacrylamide gels than enzyme incubated without ATP or with low concentrations of ATP. Comparative tryptic peptide mapping of pp60v-src incubated with low and high levels of ATP revealed that more extensive tyrosine phosphorylation of the pp60v-src polypeptide occurred at the higher concentrations of ATP. This more extensive phosphorylation was characterized by the appearance of several new phosphorylated tyrosine residues on both the amino-terminal and carboxy-terminal portions of the pp60v-src molecule. The possible consequences of these modifications on the protein kinase activity of pp60v-src, and the functional regulation of retrovirus transforming proteins in general, are discussed.