Chlamydomonas acidophila Negoro is a green algal species abundant in acidic waters (pH 2–3.5), in which inorganic carbon is present only as CO2. Previous studies have shown that aeration with CO2 increased its maximum growth rate, suggesting CO2 limitation under natural conditions. To unravel the underlying physiological mechanisms at high CO2 conditions that enables increased growth, several physiological characteristics from high- and low-CO2-acclimated cells were studied: maximum quantum yield, photosynthetic O2 evolution (Pmax), affinity constant for CO2 by photosynthesis (K0.5,p), a CO2-concentrating mechanism (CCM), cellular Rubisco content and the affinity constant of Rubisco for CO2 (K0.5,r). The results show that at high CO2 concentrations, C. acidophila had a higher K0.5,p, Pmax, maximum quantum yield, switched off its CCM and had a lower Rubisco content than at low CO2 conditions. In contrast, the K0.5,r was comparable under high and low CO2 conditions. It is calculated that the higher Pmax can already explain the increased growth rate in a high CO2 environment. From an ecophysiological point of view, the increased maximum growth rate at high CO2 will likely not be realised in the field because of other population regulating factors and should be seen as an acclimation to CO2 and not as proof for a CO2 limitation.