Expression of a clostridial [FeFe]-hydrogenase in Chlamydomonas reinhardtii prolongs photo-production of hydrogen from water splitting

The high oxygen (O-2) sensitivity of green algal [FeFe]-hydrogenases is a significant limitation for the sustained production of hydrogen gas (H-2) from photosynthetic water splitting. To address this limitation we replaced the native [FeFe]-hydrogenases with a more O-2-tolerant clostridial [FeFe]-hydrogenase CaI in Chlamydomonas reinhardtii strain D66.HYD (hydA1(-)hydA2(-)) that contains insertionally inactivated [FeFe]-hydrogenases genes. Expression and translocation of CaI in D66.HYD led to the recovery of H-2 photoproduction at similar to 20% of the rates of the wild-type parent strain D66. We show for the first time that a bacterial [FeFe]-hydrogenase can be expressed, localized and matured to a catalytically active form that couples to photosynthetic electron transport in the green alga C. reinhardtii. The lower rates of O-2 inactivation of CaI led to more sustained H-2 photoproduction when cultures were challenged with O-2 or kept under prolonged illumination at solar intensities. These results provide newinsights into the requisites for attaining photobiological H-2 production fromwater splitting using a more O-2-tolerant hydrogenase. (C) 2016 Published by Elsevier B. V.