Active recombinant human tyrosine kinase c-Yes: Expression in baculovirus system, purification, comparison to c-Src, and inhibition by a c-Src inhibitor

C-Yes is a non-receptor-type tyrosine kinase of the Src family that is most closely related to c-Src, C-Yes has been implicated in development of some human cancers. Here we report on the expression, purification, and charact erization of the active human recombinant c-Yes. A full-length human c-Yes clone has been. generated and the protein was expressed in insect Sf9 cells . Active c-Yes was purified by liquid chromatography to yield a preparation with a high specific activity (160 nmol/min/mg using an optimal Src substr ate peptide). In a comparison between human c-Yes and c-Src enzymes, relati ve phosphorylation efficiencies on nine protein and four peptide substrates were different. However, the recently described Src inhibitor CGP77675 inh ibited human c-Yes with a potency similar to that of c-Src (IC50 value of 6 .5 nM). The purified preparation of active c-Yes provides a good basis for further enzymatic characterization and for the development of c-Yes-specifi c inhibitors, (C) 2000 Academic Press.