Clostridium beijerinckii displays a soluble [FeFe]-hydrogenase/formate dehydrogenase enzyme complex that links H2 and CO2 metabolism

Abstract

The production of biofuels by bacterial fermentation receives sustained attention due to the need to develop novel circular and sustainable technologies. Clostridium beijerinckii produces both hydrogen (H2) and carbon-based biofuels acetone, butanol and ethanol (ABE solvents). H2 metabolism in C. beijerinckii is complex and mostly unexplored. Seven hydrogenase genes are contained in the genome, but their exact physiological role is unknown. Here, we report on the characterisation of a novel heterotetrameric soluble enzyme complex composed of an [FeFe]-hydrogenase component stably bound to a formate dehydrogenase subunit, which we name CbFdh/Hyd. We show that the four subunits form a stable complex that can be conveniently overexpressed and purified recombinantly. CbFdh/Hyd is highly sensitive to atmospheric oxygen and displays reversible catalytic features, including H2 evolution, H2 uptake, formate oxidation and the ability to split formate into H2 and CO2 (formate hydrogen lyase activity, FHL) as well as the opposite reaction, H2-driven CO2 reduction (HDCR). CbFdh/Hyd displays functional and spectroscopic features very similar to Fdh/Hyd complexes previously described in acetogens, suggesting that this enzyme is at the basis of the previously reported unconventional ability of C. beijerinckii to fix CO2 into acetate and butyrate. CbFdh/Hyd could also represent a key player in H2 production metabolism by degrading formate produced from the decarboxylation of pyruvate.

Study details

Article type

Original Research

Dose comparison

Kinetic parameters (KM) for H2 and formate were determined, and specific activities for different reactions (H2 evolution, H2 uptake, formate oxidation, FHL, HDCR) were compared.

Dose-dependent effect

Yes

Outcome

Positive

Findings

The study characterizes a novel soluble [FeFe]-hydrogenase/formate dehydrogenase enzyme complex (CbFdh/Hyd) from Clostridium beijerinckii, demonstrating its reversible catalytic activity in both H2 evolution and uptake, formate oxidation, and CO2 reduction. The enzyme complex is highly sensitive to oxygen and may play a key role in both H2 production and CO2 fixation in C. beijerinckii, with potential implications for biofuel production.