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The fusion of hydrogenases and photosynthetic reaction centers (RCs) has proven to be a promising strategy for the production of sustainable biofuels. Type I (iron-sulfur-containing) RCs, acting as photosensitizers, are capable of promoting electrons to a redox state that can be exploited by hydrogenases for the reduction of protons to dihydrogen (H₂). While both [FeFe] and [NiFe] hydrogenases have been used successfully, they tend to be limited due to either O₂sensitivity, binding specificity, or H₂production rates. In this study, we fuse a peripheral (stromal) subunit of Photosystem I (PS I), PsaE, to an O₂-tolerant [FeFe] hydrogenase fromClostridium beijerinckiiusing a flexible [GGS]₄linker group (CbHydA1-PsaE). We demonstrate that theCbHydA1 chimera can be synthetically activated in vitro to show bidirectional activity and that it can be quantitatively bound to a PS I variant lacking the PsaE subunit. When illuminated in an anaerobic environment, the nanoconstruct generates H₂at a rate of 84.9 ± 3.1 µmol H₂mg_chl^–1h^–1. Further, when prepared and illuminated in the presence of O₂, the nanoconstruct retains the ability to generate H₂, though at a diminished rate of 2.2 ± 0.5 µmol H₂mg_chl^–1h^–1. This demonstrates not only that PsaE is a promising scaffold for PS I-based nanoconstructs, but the use of an O₂-tolerant [FeFe] hydrogenase opens the possibility for an in vivo H₂generating system that can function in the presence of O₂.