Fractionation of carbonic anhydrase activity in Gracilaria sp. (Rhodophyta) and Enteromorpha intestinalis (Chlorophyta): changes in the extracellular activity in response to inorganic carbon levels

Abstract

The presence of different carbonic anhydrase (EC 4.2.1.1) activities has been investigated in the intertidal macroalgae Gracilaria sp. and Enteromorpha intestinalis (L.) Nees by using fractionation techniques. Activities, measured potentiometrically, were recorded for all fractions in both species, including those containing proteins associated with chloroplast membranes. In Gracilaria sp., most of the total activity was present in the soluble fraction, while similar activities were obtained for all fractions in E. intestinalis. By using inhibitors with a different capacity to enter the cell (acetazolamide and 6-ethoxyzolamide, inhibitors of external and total activity, respectively), a surface-accessible location was indicated for a high proportion of the soluble activity obtained in Gracilaria sp. In E. intestinalis, the inhibitor assays showed a substantial dependence of photosynthesis on intracellular activity. The short-term regulation of the extracellular activity in response to inorganic carbon availability was also examined in both macroalgae. Rapid repression (after 2 h) of the activity was recorded when Gracilaria sp. was transferred from limited to replete carbon conditions, while a fairly constant activity was recorded for E. intestinalis. In contrast, an increase of external activity was obtained for both macroalgae after being transferred to carbon-limited conditions, this response being more pronounced in E. intestinalis. Our results suggest the occurrence of a species-specific carbonic anhydrase system.