Direct phosphorylation of brain tyrosine hydroxylase by cyclic AMP-dependent protein kinase: Mechanism of enzyme activation

Abstract

Tyrosine hydroxylase [tyrosine monooxygenase, L-tyrosine, tetrahydropteridine:oxygen oxidoreductase (3-hydroxylating), EC 1.14.16.2] was highly purified from rat caudate nuclei. When the pure hydroxylase was phosphorylated by incubation with cylic AMP-dependent protein kinase and [32P]ATP, 32P and tyrosine hydroxylase activity were detected after polyacrylamide gel electrophoresis in a single protein band. After sodium dodecyl sulfate gel electrophoresis, 32P was detected only in a probable active subunit of tyrosine hydroxylase of molecular weight 62,000. Phosphorylation of the hydroxylase increased its activity by 2-fold, and was associated with an increase in V(m) without any change in K(m) for either substrate or cofactor. It is proposed that the pool of native tyrosine hydroxylase is composed of a mixture of enzyme molecules in both active and probably inactive forms, that the active form is phosphorylated, and that phosphorylation produces an active form of the enzyme at the expense of an inactive one.