Carbonic Anhydrase and the Uptake of Inorganic Carbon by Synechococcus sp. (UTEX-2380) 1

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We report the changes in the concentrations and 18O contents of extracellular CO2 and HCO3− in suspensions of Synechococcus sp. (UTEX 2380) using membrane inlet mass spectrometry. This marine cyanobacterium is known to have an active uptake mechanism for inorganic carbon. Measuring 18O exchange between CO2 and water, we have found the intracellular carbonic anhydrase activity to be equivalent to 20 times the uncatalyzed CO2 hydration rate in different samples of cells that were grown on bubbled air (low-CO2 conditions). This activity was only weakly inhibited by ethoxzolamide with an I50 near 7 to 10 micromolar in lysed cell suspensions. We have shown that even with CO2-starved cells there is considerable generation of CO2 from intracellular stores, a factor that can cause errors in measurement of net CO2 uptake unless accounted for. It was demonstrated that use of 13C-labeled inorganic carbon outside the cell can correct for such errors in mass spectrometric measurement. Oxygen-18 depletion experiments show that in the light, CO2 readily passes across the cell membrane to the sites of intracellular carbonic anhydrase. Although HCO3− was readily taken up by the cells, these experiments shown that there is no significant efflux of HCO3− from Synechococcus.

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