Alterations in nitrogen metabolism caused by heavy metals in the acid-tolerant microalga Coccomyxa onubensis

Abstract

The microalga Coccomyxa onubensis is an extremophile microorganism with a unique ecosystem (Río Tinto, Huelva, Spain) that contains high amounts of contaminants, including heavy metals, sulphates, and nitrates, in acidic environments (pH 2.5). The current work presents an evaluation of the capacity of Coccomyxa onubensis to assimilate different nitrogen sources under Cu 2+ , Cd 2+ , AsO 3 3− , AsO 4 3− and Hg 2+ stress, and the metabolic implications of these stressors. The results showed that ammonium consumption was less affected than nitrate consumption when microalgae were cultivated with heavy metals (except cadmium). The activities of enzymes involved in nitrogen metabolism, such as nitrite reductase (NiR; EC:1.7.7.1), glutamine synthetase (GS; EC:6.3.2.1) and glutamate dehydrogenase (GDH; EC:1.4.1.2) were characterised to determine the Michaelis-Menten constant ( K m ) and optimal temperature and pH values, being 45, 40 and 60 °C and pH values of 7.5, 6.0 and 9.0 for NiR, GS, and GDH, respectively. The effects of different heavy metals on these enzymes were assessed at multiple levels, and the results showed that the enzymatic activity of NiR was downregulated, specially under copper stress, maintaining 23 % of control NiR activity at 2 mM Cu 2+ . The enzymatic activity of GS was upregulated at low concentrations under cadmium and mercury stress (115–120 % of control cultures GS activity at 25 μM Cd 2+ and 50 nM Hg 2+ , respectively) and downregulated at high concentrations of these elements. GDH activity was upregulated in the presence of Cu 2+ , Cd 2+ , and Hg 2+ , with increases up to 192, 155 and 154 % at 1 mM Cu 2+ , 300 μM Cd 2+ , and 250 nM Hg 2+ , respectively. These results provide a better explanation of the effects of heavy metal stress on N metabolism in Coccomyxa onubensis , which may be used as a model eukaryotic organism of the Tinto River acidophilic ecosystem . • C. onubensis tolerates high amounts of heavy metal in cultures with a pH 2.5. • Under heavy metal stress, nitrate consumption was less altered than ammonium one. • N metabolism enzymes were characterised for the microalga C. onubensis. • NiR and GS enzymatic activities were downregulated under heavy metal stress. • GDH was upregulated under heavy metal stress, likely for L-glutamate synthesis.

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