Tyrosine Hydroxylase Inactivation Following cAMP‐Dependent Phosphorylation Activation

Abstract

Tyrosine hydroxylase, the rate-limiting enzyme in catecholamine biosynthesis, is activated following phosphorylation by the cAMP-dependent protein kinase (largely by decreasing the Km of the enzyme for its pterin co-substrate). Following its phosphorylation activation in rat striatal homogenates, we find that tyrosine hydroxylase is inactivated by two distinct processes. Because cAMP is hydrolyzed in crude extracts by a phosphodiesterase, cAMP-dependent protein kinase activity declines following a single addition of cAMP. When tyrosine hydroxylase is activated under these transient phosphorylation conditions, inactivation is accompanied by a reversion of the activated kinetic form (low apparent Km for pterin co-substrate, less than or equal to 0.2 mM) to the kinetic form characteristic of the untreated enzyme (high apparent Km, greater than or equal to 1.0 mM). This inactivation is readily reversed by the subsequent addition of cAMP. When striatal tyrosine hydroxylase is activated under constant phosphorylation conditions (incubated with purified cAMP-dependent protein kinase catalytic subunit), however, it is also inactivated. This second inactivation process is irreversible and is characterized kinetically by a decreasing apparent Vmax with no change in the low apparent Km for pterin co-substrate (0.2 mM). The latter inactivation process is greatly attenuated by gel filtration which resolves a low-molecular-weight inactivating factor(s) from the tyrosine hydroxylase. These results are consistent with a regulatory mechanism for tyrosine hydroxylase involving two processes: in the first case, reversible phosphorylation and dephosphorylation and, in the second case, an irreversible loss of activity of the phosphorylated form of tyrosine hydroxylase.

Biochemical Conservation of Recombinant Drosophila Tyrosine Hydroxylase with its Mammalian Cognates

“…generated DTH results in diminished enzymatic activity with increasing incubation time, consistent with observations from studies with mammalian TH that suggest the phosphorylated form is less stable (Lazar et al, 1981;Vrana and Roskoski, 1983). Our results show that PKA phosphorylation of DA-bound DTH activates the enzyme, but that PKA phosphorylation of protein without bound DA results in reduced activity.…”

Protein Kinase Cδ Negatively Regulates Tyrosine Hydroxylase Activity and Dopamine Synthesis by Enhancing Protein Phosphatase-2A Activity in Dopaminergic Neurons

“…Previous studies have demonstrated that PP2A is a major serine phosphatase that mediates the dephosphorylation of TH, in particular TH-ser40, resulting in the inactivation of TH (Vrana and Roskoski, 1983;Haavik et al, 1989). Because both of these proteins negatively regulate TH, we hypothesize that PKC␦ may work through PP2A to reduce dephosphorylation of TH-ser40 and TH activity.…”

Normal and Physio-Pathological Striatal Dopamine Homeostasis

“…Therefore increased TH activity, which is often reported after partial lesion of the DA pathway could be, at least in part, a consequence of reduced intraterminal DA pools. The simultaneous reduction in the amount of tissue TH protein may occur in one of two ways: (1) a reduction in the number of DA terminals, or (2) increased protein turnover associated with the increased catalytic activity (Vrana and Roskoski, 1983;Lavergne et al, 1994).…”