Phosphorylation of Ser(19) alters the conformation of tyrosine hydroxylaseto increase the rate of phosphorylation of Ser(40)

The effect of phosphorylation on the shape of tyrosine hydroxylase (TH) was studied directly using gel filtration and indirectly using electrospray io nization mass spectrometry. Phosphorylation of Ser(19) and Ser(40) produced a TH molecule with a more open conformation than the non-phosphorylated fo rm. The conformational effect of Ser(19) phosphorylation is less pronounced than that of the Ser(40) phosphorylation. The effect of Ser(19) and Ser(40 ) phosphorylation appears to be additive. Binding of dopamine produced a mo re compact form when compared with the non-dopamine-bound TH. The interdepe ndence of Ser(19) and Ser(40) phosphorylation was probed using electrospray ionization mass spectrometry. The rate constants for the phosphorylation o f Ser(19) and Ser(40) were determined by electrospray ionization mass spect rometry using a consecutive reaction model. The rate constant for the phosp horylation of Ser(40) is similar to2 to 3-fold higher if Ser(19) is already phosphorylated. These results suggest that phosphorylation of Ser(19) alte rs the conformation of tyrosine hydroxylase to allow increased accessibilit y of Ser(40) to kinases.