Background: Chlamydomonas contains six chloroplast ferredoxins (FDXs) whose function is still unclear. Results: A global FDX interactome was obtained where FDX1 has a predominant role and is the most relevant electron donor to FNR1 and HYDA1. Conclusion: FDXs have distinct but also overlapping function. Significance: We discovered new FDX interaction partners and specific roles for each FDX isoform.

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 D66ΔHYD (hydA1-hydA2-) that contains insertionally inactivated [FeFe]-hydrogenases genes. Expression and translocation of CaI in D66ΔHYD led to the recovery of H 2 photoproduction at ~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 new insights into the requisites for attaining photobiological H 2 production from water splitting using a more O 2-tolerant hydrogenase.

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