CHARACTERIZATION OF INORGANIC CARBON FLUXES, CARBONIC ANHYDRASE(S) AND RIBULOSE-1,5-BIPHOSPHATE CARBOXYLASE-OXYGENASE IN THE GREEN UNICELLULAR ALGA COCCOMYXA - COMPARISONS WITH LOW-CO2 CELLS OF CHLAMYDOMONAS-REINHARDTII

Processes involved in the uptake and fixation of dissolved inorganic c arbon (DIG) were characterised for Coccomyxa, the green algal primary photobiont of the lichen Peltigera aphthosa and compared with the free -living alga Chlamydomonas reinhardtii Dangeard (WT cc 125(+)). A mass -spectrometer disequilibrium technique was used to quantify fluxes of both HCO3- and CO2 in the two algae, while activities of carbonic anhy drases (CAs) were examined in intact cells by measuring O-18 exchange from doubly labelled CO2 ((COO)-C-13-O-18-O-18) to water and by using CA inhibitors. The CO2-fixation kinetics of intact Coccomyxa cells wer e also compared with the carboxylation efficiency of its isolated and purified primary carboxylating enzyme, ribulose-1,5-bisphosphate carbo xylase-oxygenase (Rubisco). The two algae were found to be significant ly different in their modes of acquiring CO2 for photosynthesis. Chlam ydomonas was able to actively transport both HCO3- and CO2 from the ex ternal medium, while Coccomyxa clearly favoured CO2 as a substrate. Bo th algae were found to possess external as well as internal CAs, but t he relative amounts of these as well as their overall significance for the functioning of photosynthesis differed. In Coccomyxa, the interna l CA activity was significantly higher than in Chlamydomonas and also predominated over the external activity. In Chlamydomonas, both transp ort and fixation of DIC were severely inhibited by ethoxyzolamide, an inhibitor of external and internal CAs as well as the DIG-transporting system, while this inhibitor only caused a minor inhibition of photos ynthesis in Coccomyxa. These results thus give strong support for earl ier indirect observations of the absence of a CO2-concentrating mechan ism in Coccomyxa. In addition, Coccomyxa was found to possess a Rubisc o with a higher carboxylation efficiency than Chlamydomonas, having a Km(CO2) of 12 +/- 3 mu M CO2 and a CO2/O-2 specificity factor (S-c/o) of 83 +/- 2, and it may hence be concluded that the absence of the CO2 -concentrating mechanism is positively correlated with a more efficien t Rubisco in this alga.