Thin cell layer cultures of Chlamydomonas reinhardtii L159I-N230Y, pgrl1 and pgr5 mutants perform enhanced hydrogen production at sunlight intensity.

Photobiological hydrogen (H) production is a promising renewable energy source. HydA hydrogenases of green algae are efficient but O-sensitive and compete for electrons with CO-fixation. Recently, we established a photoautotrophic H production system based on anaerobic induction, where the Calvin-Benson cycle is inactive and O scavenged by an absorbent. Here, we employed thin layer cultures, resulting in a three-fold increase in H production relative to bulk CC-124 cultures (50 µg chlorophyll/ml, 350 µmol photons m s). Productivity was maintained when increasing the light intensity to 1000 µmol photons ms and the cell density to 150 µg chlorophyll/ml. Remarkably, the L159I-N230Y photosystem II mutant and the pgrl1 photosystem I cyclic electron transport mutant produced 50% more H than CC-124, while the pgr5 mutant generated 250% more (1.2 ml H/ml culture in six days). The photosynthetic apparatus of the pgr5 mutant and its in vitro HydA activity remained remarkably stable.