A Single Cyclohexadienyl Dehydrogenase Specifies the Prephenate Dehydrogenase and Arogenate Dehydrogenase Components of the Dual Pathways to L-Tyrosine in Pseudomonas aeruginosa*

Dual biosynthetic pathways diverge from prephenate to L-tyrosine in Pseudomonas aeruginosa, with 4-hydroxyphenylpyruvate and L-arogenate being the unique intermediates of these pathways. Prephenate dehydrogenase and arogenate dehydrogenase activities could not be separated throughout fractionation steps yielding a purification of more than 200-fold, and the ratio of activities was constant throughout purification. Thus, the enzyme is a cyclohexadienyl dehydrogenase. The native enzyme has a molecular weight of 150,000 and is a hexamer made up of identical 25,500 subunits. The enzyme is specific for NAD+ as an electron acceptor, and identical K,,, values of 0.25 mM were obtained for NAD+, regardless of whether activity was assayed as prephenate dehydrogenase or as arogenate dehydrogenase. K,,, values of 0.07 mM and 0.17 mM were calculated for prephenate and Larogenate, respectively. Inhibition by L-tyrosine was noncompetitive with respect to NAD’, but was strictly competitive with respect to either prephenate or Larogenate. With cyclohexadiene as variable substrate, similar Ki values for L-tyrosine of 0.06 mM (prephenate) and 0.05 mM (L-arogenate) were obtained. With NAD+ as the variable substrate, similar Ki values for L-tyrosine of 0.26 mM (prephenate) and 0.28 mM (Larogenate), respectively, were calculated. This is the first characterization of a purified, monofunctional cyclohexadienyl dehydrogenase.