Soluble, Membrane-bound, and Detergent-solubilized Rat Striatal Tyrosine Hydroxylase

Soluble striatal tyrosine hydroxylase undergoes a dramatic increase in affinity for the synthetic cofactor 2-amino-4-hydroxy-6,7-dimethyl-5,6,7,8-tetrahydropteridine (DMPH4) from 1.5 mm above pH 6 to 0.25 mm below pH 5.7. In the presence of 0.075% Triton X-100, the Km for DMPH4 below pH 5.7 is further reduced to 0.1 mm. The presence of the specific mucopolysaccharide, heparin, which exhibits a Ka of near 10 µg per ml independent of pH, and cooperativity of binding, totally abolishes the pH-dependent change, and alters the Km for DMPH4 to 0.1 mm. Under all conditions, the soluble enzyme exhibits a Km for tyrosine near 56 µm. Solubilization of membrane-bound tyrosine hydroxylase with Triton X-100 yields a form of the enzyme identical with the soluble enzyme in kinetic properties as well as on sucrose density gradients. On the other hand, membrane-bound tyrosine hydroxylase exhibits a Km for DMPH4 near 0.1 mm independent of pH. Further, the membrane-bound enzyme has a Km for tyrosine of 8 to 10 µm. The data are consistent with a highly cooperative conformational transition as the pH is raised from 5.7 to 6.0, which is prevented by the presence of heparin, or by the binding of tyrosine hydroxylase to membrane components. Further, the observation that synaptic membrane-bound tyrosine hydroxylase exhibits a decreased Km for both tryosine and DMPH4 could provide an alternative mechanism by which newly synthesized transmitter may be preferentially released.