A single cyclohexadienyl dehydratase specifies the prephenate dehydratase and arogenate dehydratase components of one of two independent pathways to L-phenylalanine in Erwinia herbicola.

ABSTRACT

Dual biosynthetic pathways diverge from prephenate to L-phenylalanine in Erwinia herbicola, the unique intermediates of these pathways being phenylpyruvate and L-arogenate. After separation from the bifunctional P-protein (one component of which has prephenate dehydratase activity), the remaining prephenate dehydratase activity could not be separated from arogenate dehydratase activity throughout fractionation steps yielding a purification of more than 1200-fold. The ratio of activities was constant after removal of the P-protein, and the two dehydratase activities were stable during purification. Hence, the enzyme is a cyclohexadienyl dehydratase. The native enzyme has a molecular mass of 73 kDa and is a tetramer made up of identical 18-kDa subunits. Km values of 0.17 mM and 0.09 mM were calculated for prephenate and L-arogenate, respectively. L-Arogenate inhibited prephenate dehydratase competitively with respect to prephenate, whereas prephenate inhibited arogenate dehydratase competitively with respect to L-arogenate. Thus, the enzyme has a common catalytic site for utilization of prephenate or L-arogenate as alternative substrates. This is the first characterization of a purified monofunctional cyclohexadienyl dehydratase.