Abstract:
Nitrogen-deprived cells of the marine diatom Phaeodactylum tricornutum possessed extracellular L-amino acid oxidase (EC 1.4.3.2) activity. No enzyme activity was detectable in nitrogen-replete cells. When nitrogen-deprived cells were provided with 100 µM alanine, the amino acid was metabolised, with both pyruvate and H^sub 2^O^sub 2^ accumulating stoichiometrically in the medium; this accumulation was largely prevented if the cells were preincubated with the membrane-impermeable protease Proteinase K. Following the transfer of nitrogen-replete cells to nitrogen-free medium, development of enzyme activity lagged, with little or no enzyme activity detectable until 4 h after the onset of nitrogen deprivation. This lag was followed by rapid development of enzyme activity during the ensuing 20 h. In addition to alanine, methionine, methionine sulfoximine, leucine, glutamate and valine, at the same initial concentration of 100 µM, were exclusively metabolised by an amino acid oxidase. Histidine was partially metabolised by an amino acid oxidase, whereas arginine and lysine were transported into the cell with little or no metabolism by an amino acid oxidase. Neither serine nor proline was used. In general, the neutral nonpolar and acidic polar amino acids were used via an extracellular amino acid oxidase, whereas the basic polar amino acids were transported. L-Amino acid oxidase activity was inhibited in darkness and in light in the presence of the photosynthetic inhibitor DCMU (3-[3,4-dichlorophenyl]-1,1-dimethylurea). In addition, enzyme activity decreased on addition of bicarbonate to cells in light. Together these observations suggest that extracellular L-amino acid oxidase activity was inhibited by dissolved inorganic carbon.
