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Analysis of the reaction of dopa decarboxylase (DDC) with L-dopa reveals that loss of decarboxylase activity with time is observed at enzyme concentrations approximately equal to the binding constant, Kd, of the enzyme for pyridoxal 5ƀ-phosphate (PLP). Instead, at enzyme concentrations higher than Kd the course of product formation proceeds linearly until complete consumption of the substrate. Evidence is provided that under both experimental conditions no pyridoxamine 5ƀ-phosphate (PMP) is formed during the reaction and that dissociation of coenzyme occurs at low enzyme concentration, leading to the formation of a PLP-L-dopa Pictet–Spengler cyclic adduct. Taken together, these results indicate that decarboxylation-dependent transamination does not accompany the decarboxylation of L-dopa proposed previously [O'Leary and Baughn (1977) J. Biol. Chem. 252, 7168–7173]. Nevertheless, when the reaction of DDC with L-dopa is studied under anaerobic conditions at an enzyme concentration higher than Kd, we observe that (1) the enzyme is gradually inactivated and inactivation is associated with PMP formation and (2) the initial velocity of decarboxylation is approximately half of that in the presence of O2. Similar behaviour is observed by comparing the reaction with L-5-hydroxytryptophan occurring in aerobiosis or in anaerobiosis. Therefore the reaction of DDC with L-aromatic amino acids seems to be under O2 control. In contrast, the reactivity of the enzyme with D-aromatic amino acids does not change in the presence or absence of O2. These and other results, together with previous results on the effect exerted by O2 on reaction specificity of DDC towards aromatic amines [Bertoldi, Frigeri, Paci and Borri Voltattorni (1999) J. Biol. Chem. 274, 5514–5521], suggest a productive effect of O2 on an intermediate complex of the reaction of the enzyme with L-aromatic amino acids or aromatic amines.

Ornithine decarboxylase (ODC) from Lactobacillus 30a catalyses the cleavage of alpha-methylornithine into ammonia and 2-methyl-1-pyrroline; glutamate decarboxylase (GAD) from Escherichia coli catalyses the cleavage of alpha-methylglutamate into ammonia and laevulinic acid. In our analyses, 2-methyl-1-pyrroline and laevulinic acid were identified by HPLC and mass spectroscopic analysis, and ammonia was identified by means of glutamate dehydrogenase. Molecular oxygen was consumed during these reactions in a 1:2 molar ratio with respect to the products. The catalytic efficiencies (k(cat)/K(m)) of the reactions catalysed by ODC and GAD were determined as 12500 and 9163 M(-1).min(-1) respectively. When the reactions were performed under anaerobic conditions, no ammonia, 2-methyl-1-pyrroline or laevulinic acid was produced to a significant extent. The formation of ammonia and O(2) consumption (in a 1:2 molar ratio with respect to ammonia) were also detected during the reaction of ODC and GAD with putrescine and gamma-aminobutyrate respectively. Taken together, these findings clearly indicate that ODC and GAD catalyse an oxidative deamination of their decarboxylation products, a reaction similar to that catalysed by dopa decarboxylase (DDC) with alpha-methyldopa [Bertoldi, Dominici, Moore, Maras and Borri Voltattorni (1998) Biochemistry 37, 6552-6561]. Furthermore, this reaction was accompanied by a decarboxylation-dependent transamination occurring for GAD, DDC and ODC with a frequency of approx. 0.24%, 1% and 9% respectively compared with that of oxidative deamination.