Reaction of dopa decarboxylase with L-aromatic amino acids under aerobic and anaerobic conditions.
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, K(d), of the enzyme for pyridoxal 5'-phosphate (PLP). Instead, at enzyme concentrations higher than K(d) the course of product formation proceeds linearly until complete consumption of the substrate. Evidence is provided that under both experimental conditions no pyridoxamine 5'-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 K(d), 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 O(2). 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 O(2) control. In contrast, the reactivity of the enzyme with L-aromatic amino acids does not change in the presence or absence of O(2). These and other results, together with previous results on the effect exerted by O(2) 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 O(2) on an intermediate complex of the reaction of the enzyme with L-aromatic amino acids or aromatic amines.
Bertoldi
M
M
Dipartimento di Scienze Neurologiche e della Visione, Sezione di Chimica Biologica, Facoltà di Medicina e Chirurgia, Università degli Studi di Verona, Strada Le Grazie, 8, 37134 Verona, Italy.
Borri Voltattorni
C
C
D006916:5-Hydroxytryptophan / Q000378:metabolism; D000596:Amino Acids / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D004296:Dopa Decarboxylase / Q000378:metabolism*; D006868:Hydrolysis; D007980:Levodopa / Q000378:metabolism; D009955:Ornithine Decarboxylase / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism; D011732:Pyridoxal Phosphate / Q000378:metabolism
D000596:Amino Acids; D007980:Levodopa; D011732:Pyridoxal Phosphate; D006916:5-Hydroxytryptophan; D004296:Dopa Decarboxylase; D009955:Ornithine Decarboxylase; D010100:Oxygen