Autophosphorylation of the insulin-like growth factor I receptor cytoplasmic domain

The cytoplasmic domain of the beta subunit of the insulin-like growth facto r I receptor (amino acids 936-1337) was overexpressed in Sf9 insect cells u sing a baculovirus expression system, and the g-His tagged receptor was pur ified by metal-affinity chromatography. Autophosphorylation of the receptor was concentration dependent, consistent with a trans phosphorylation mecha nism, Phosphoamino acid analysis of the autophosphorylated receptor showed predominantly phosphotyrosine, but phosphoserine and phosphothreonine were also present. However, when the receptor was further purified by gel filtra tion on Sephadex G-100 and then autophosphorylated, phosphoaminoacid analys is showed only phosphotyrosine, We conclude that the IGF-I receptor tyrosin e kinase is not a dual-specificity kinase and that autophosphorylation of t he beta subunit is by a trans mechanism.