EXTRACELLULAR AND INTRACELLULAR CARBONIC-ANHYDRASE IN RELATION TO CULTURE AGE IN A HIGH-CALCIFYING STRAIN OF EMILIANIA-HUXLEYI LOHMANN

The relationships between extra- and intra-cellular carbonic anhydrase, calcification rate, utilization of dissolved inorganic carbon (DIC) and culture age were investigated in a high-calcifying strain of Emiliania huxleyi. The detection of carbonic anhydrase was dependent on culture age; neither the extra-cellular activity of intact cells nor activity in crude homogenates was detected until the stationary phase. By the stationary phase DIC in the medium was totally depleted and the calcification rate had decreased by 60%. Extra- and intra-cellular carbonic anhydrase was stimulated by Na+ but not Cl ions. The isolation of intact organelles by isopycnic gradient centrifugation from exponential and stationary phase cultures of high calcifying cells showed high carbonic anhydrase activity in the chloroplast fraction but carbonic anhydrase was not detected in low-calcifying cells even after gradient centrifugation. Although 50 mu M ethoxyzolamide was an effective inhibitor of carbonic anhydrase activity in vitro, photosynthetic (CO2)-C-14 fixation was only inhibited 30%, calcification rate 10% and the internal inorganic carbon pool unaffected in intact cells. It is proposed that chloroplast carbonic anhydrase maintains the steady state flux of CO2 from the chloroplast envelope to Rubisco, much of the CO2 arising by release from HCO3- in the cytosol, required to maintain cytoplasmic pH near neutrality in high-calcifying cells.