Autotrophic hydrogen photoproduction by operation of carbon-concentrating mechanism in Chlamydomonas reinhardtii under sulfur deprivation condition

Under autotrophic conditions, starch plays an important role in establishing anoxic conditions during PSII-dependent hydrogen (H₂) photoproduction in microalgae. This is because starch is the sole organic substrate during respiratory consumption of internal oxygen (O₂) from PSII-dependent direct pathway. Herein, we propose a novel approach to further facilitate the internal starch synthesis of Chlamydomonas reinhardtii through the operation of carbon-concentrating mechanism (CCM) along with a two-stage process based on sulfur (S) deprivation, thereby resulting in enhanced anaerobic capacity during PSII-dependent H₂ photoproduction. When CCM-induced cells were exposed to high levels of carbon dioxide (CO₂) (5%, v/v) with S deprivation, internal levels of starch were significantly elevated by retaining a functional CCM with the boosted photosynthetic activity during 24h of O₂ evolution phase (I) of S deprivation. Consequently, during H₂ production phase of S deprivation at irradiance of 50μEm^-2s^-1, the concentrations of starch and H₂ in CCM-induced cells were remarkably enhanced by 65.0% and 218.9% compared to that of CCM-uninduced cells, respectively. The treatment of low-CO₂-driven CCM induction prior to S deprivation is a cost-effective and energy-efficient strategy that significantly improves the solar-driven H₂ production by microalgae; this is particularly realizable in an industrial scale.