title: Use of tyrosine-containing polymers to characterize the substrate specificity of insulin and other hormone-stimulated tyrosine kinases.
abstract: Synthetic copolymers containing tyrosine residues were used to characterize the substrate specificity of the insulin receptor kinase and compare it to tyrosine kinases stimulated by epidermal growth factor, insulin-like growth factor-1 and phorbol ester. In partially purified receptor preparations from eight different tissues insulin best stimulated (highest V) phosphorylation of a random copolymer composed of glutamic and tyrosine residues at a 4:1 ratio (Glu/Tyr, 4:1). The insulin-stimulated phosphorylation of this polymer was highly significant also in receptor preparations from fresh human monocytes, where insulin binding and autophosphorylation were difficult to detect. Other tyrosine-containing polymers Ala/Glu/Lys/Tyr (6:2:5:1) and Glu/Ala/Tyr (6:3:1) were also phosphorylated by the insulin-stimulated kinase but to a lower extent. A tyrosine kinase stimulated by insulin-like growth factor-1, and one stimulated by phorbol ester also best phosphorylated the polymer Glu/Tyr (4:1). The three kinases differed only in their capability to phosphorylate Glu/Ala/Tyr (6:3:1) or Ala/Glu/Lys/Tyr (6:2:5:1). Glu/Tyr (4:1) was a poor substrate for the epidermal growth factor receptor kinase which best phosphorylated the polymer Glu/Ala/Tyr (6:3:1). Three additional polymers: Glu/Tyr (1:1), Glu/Ala/Tyr (1:1:1), and Lys/Tyr (1:1) failed to serve as substrates for all four tyrosine kinases tested. Taken together these findings suggest that. Hormone-sensitive tyrosine kinases have similar yet distinct substrate specificity and are likely to phosphorylate their native substrates on tyrosines adjacent to acidic (glutamic) residues. Tyrosine-containing polymer substrates are highly sensitive and convenient tools to study (hormone-sensitive) tyrosine kinases whose native substrates are unknown or present at low concentrations.
authors.forename: Y
authors.initials: Y
authors.lastname: Zick
authors.forename: G
authors.initials: G
authors.lastname: Grunberger
authors.forename: R W
authors.initials: RW
authors.lastname: Rees-Jones
authors.forename: R J
authors.initials: RJ
authors.lastname: Comi
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D004789:Enzyme Activation; D006728:Hormones / Q000502:physiology*; D006801:Humans; D007328:Insulin / Q000502:physiology*; D008214:Lymphocytes / Q000201:enzymology; D051379:Mice; D010766:Phosphorylation; D011108:Polymers / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases; D051381:Rats; D011972:Receptor, Insulin; D013379:Substrate Specificity; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000378:metabolism
chemical_list: D006728:Hormones; D007328:Insulin; D011108:Polymers; D014443:Tyrosine; D011494:Protein Kinases; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin