Identification of Zn²⁺-dependent and Mg²⁺/Triton X-100-dependent tyrosine protein kinases in ethylenediaminetetraacetate-treated P₂ membrane fraction of monkey brain basal ganglia

Abstract

Tyrosine phosphorylation of a 55- and 60-kDa protein was observed when EDTA-treated P₂ membrane fraction from monkey basal ganglia was incubated with [γ-³²P]-ATP in the presence of Zn²⁺. Other metal ions were less effective in this phosphorylation. The effect of Zn²⁺ did not appear to be due to its inhibition of a tyrosine phosphatase. In the presence of Mg²⁺/Triton X-100 instead of Zn²⁺, phosphorylation on tyrosine residues of a 17-kDa protein and the external substrate poly(Glu, Tyr) 4:1 copolymer was observed. Both Mg²⁺ and Triton X-100 were essential for this and Zn²⁺ inhibited both of these phosphorylations. Convincing evidence for the existence of Zn²⁺-dependent and Mg²⁺/Triton X-100-dependent tyrosine protein kinases was obtained when the two kinases could be separated by extraction of the membranes by Triton X-100. The Zn²⁺-dependent phosphorylation was present exclusively in the Triton-solubilized supernatant whereas the Mg²⁺/Triton X-100-dependent phosphorylation was found associated with the Triton-insoluble membrane fractions. Externally added histone could also be phosphorylated on tyrosine residues in a Zn²⁺- or Mg²⁺/Triton X-100-dependent manner by the supernatant or membrane fraction, respectively.