The
development of renewable and sustainable biofuels to cover the future
energy demand is one of the most challenging issues of our time.
Biohydrogen, produced by photosynthetic microorganisms, has the
potential to become a green biofuel and energy carrier for the future
sustainable world, since it provides energy without CO₂
emission. The recent development of two alternative protocols to induce
hydrogen photoproduction in green algae enables the function of the O₂-sensitive [FeFe]-hydrogenases, located at the acceptor side of photosystem I, to produce H₂ for several days. These protocols prevent carbon fixation and redirect electrons toward H₂ production. In the present work, we employed these protocols to a knockout Chlamydomonas reinhardtii mutant lacking flavodiiron proteins (FDPs), thus removing another possible electron competitor with H₂ production.

The deletion of the FDP electron sink resulted in the enhancement of H₂ photoproduction relative to wild-type C. reinhardtii. Additionally, the lack of FDPs leads to a more effective obstruction of carbon fixation even under elongated light pulses.

We
demonstrated that the rather simple adjustment of cultivation
conditions together with genetic manipulation of alternative electron
pathways of photosynthesis results in efficient re-routing of electrons
toward H₂ photoproduction. Furthermore, the introduction of a short recovery phase by regular switching from H₂ photoproduction to biomass accumulation phase allows to maintain cell fitness and use photosynthetic cells as long-term H₂-producing biocatalysts.