The oxidative decarboxylation of ornithine by extracts of higher plants

Abstract

Extracts of wheat leaves catalysed the oxidative decarboxylation of ornithine in the presence of pyridoxal phosphate, with the production of 4-aminobutanamide and 4-aminobutyric acid. In similar experiments, putrescine was not converted to the amide or acid suggesting it is not an intermediate. The optimum concentration of pyridoxal phosphate was 1 mM, and activity declined on increasing the concentration to 10 mM. The K m for ornithine was 10 mM. The activity was found principally in the particulate fraction. Both the particulate and the soluble fractions showed two pH optima, at ca pH 9 and 10.5. Two pH optima were also found for ornithine decarboxylation in the particulate fraction of oat leaves: one at pH 6.5-7 and another at pH 9–9.5. Activity was lost on heating for 1 hr at 100°, though discontinuities in the thermal denaturation curve indicated a heterogeneous system. d- and l-Ornithine and l- lysine acted as substrates. 2,3-Diaminopropionic acid and 2,4-diaminobutyric acid were oxidized to ninhydrin-positive compounds which were presumably respectively 2-aminoacetamide and 3-aminopropanamide. Aminoguanidine (0.25 mM inhibited ornithine decarboxylation by 30–40%, while difluoromethylornithine at 1 mM showed no inhibition. Dithiothreitol, dithioerythritol, cysteine, mercaptoethanol and glutathione inhibited the oxidation at 1 and 5 mM. Particulate preparations of barley, sugar beet and rape leaves also oxidized ornithine to 4-aminobutanamide and 4-aminobutyric acid. Gel filtration on Sephadex G-25 showed that the M r of the major component causing oxidative decarboxylation of ornithine present in the wheat leaf supernatant was ca 4000. However another less active fraction was found to have a M r of > 5000. Mn 2+ ions at μM concentrations catalyse a similar decarboxylation of ornithine in the presence of pyridoxal phosphate with the production of 4-aminobutanamide, and may be responsible for the activity in the plant extracts. Mn 2+ ions are known to associate with nucleic acids and may therefore simulate a complex catalyst of relatively large M r . At high pH, Mn 2+ ions exist as the unstable hydroxide which can be readily oxidized, explaining the apparent thermal denaturation, which appears to be accelerated in the presence of the plant extracts.