Action at a distance: The remarkable coupling of CO uptake to electron transfer in specialized cyanobacterial NDH-1 complexes.

Cyanobacteria achieve highly efficient photosynthesis using a CO-concentrating mechanism relying on specialized Type I (NDH-1) complexes. Among these, NDH-1 and NDH-1 catalyze redox-coupled hydration of CO to bicarbonate, supporting carbon fixation in carboxysomes. The mechanism of coupling electron transfer to CO-hydration by these variant NDH-1 complexes remains unknown. We engineered a PCC7942 strain that expresses exclusively the high flux/low affinity NDH-1 complex, enabling the observation of the coupling of CO hydration to cyclic electron flow in isolation from the other NDH-1 isoforms normally present in cells. We found that inhibition of the CupB protein by the carbonic anhydrase inhibitor ethoxzolamide (EZ) suppressed CO uptake, slowed photosystem I rereduction, and abolished proton pumping as probed by acridine orange fluorescence. These effects were absent in strains lacking Cup proteins, confirming specificity. The results demonstrate that CO hydration and electron transfer through NDH-1 are tightly coupled via proton translocation across the thylakoid membrane. These findings provide direct evidence for the bidirectional interaction in bioenergetic coupling between the plastoquinone reduction and the CO uptake at the distal Zn-site over a span of ~150 Å and support a proton-removal hypothesis involving the proton transfer pathways from the Zn-site of CO hydration to an energetically coupled proton loading site evolutionarily repurposed from the ancestral proton pumping mechanism to enable energetic CO uptake.