Cyanobacteria, algae, and plants oxidize water to the O2 we breathe, and consume CO2 during the synthesis of biomass. Although these vital processes are functionally and structurally well separated in photosynthetic organisms, there is a long-debated role for CO2/HCO3(-) in water oxidation. Using membrane-inlet mass spectrometry we demonstrate that HCO3(-) acts as a mobile proton acceptor that helps to transport the protons produced inside of photosystem II by water oxidation out into the chloroplast's lumen, resulting in a light-driven production of O2 and CO2. Depletion of HCO3(-) from the media leads, in the absence of added buffers, to a reversible down-regulation of O2 production by about 20%. These findings add a previously unidentified component to the regulatory network of oxygenic photosynthesis and conclude the more than 50-y-long quest for the function of CO2/HCO3(-) in photosynthetic water oxidation.