O₂ sensitivity and H₂ production activity of hydrogenases—A review

Abstract

Hydrogenases are metalloproteins capable of catalyzing the interconversion between molecular hydrogen and protons and electrons. The iron–sulfur clusters within the enzyme enable rapid relay of electrons which are either consumed or generated at the active site. Their unparalleled catalytic efficiency has attracted attention, especially for potential use in H2 production and/or fuel cell technologies. However, there are limitations to using hydrogenases, especially due to their high O2 sensitivity. The subclass, called [FeFe] hydrogenases, are particularly more vulnerable to O2 but proficient in H2 production. In this review, we provide an overview of mechanistic and protein engineering studies focused on understanding and enhancing O2 tolerance of the enzyme. The emphasis is on ongoing studies that attempt to overcome O2 sensitivity of the enzyme while it catalyzes H2 production in an aerobic environment. We also discuss pioneering attempts to utilize the enzyme in biological H2 production and other industrial processes, as well as our own perspective on future applications.

Optimization of Culture Conditions for Oxygen-Tolerant Regulatory [NiFe]-Hydrogenase Production from Ralstonia eutropha H16 in Escherichia coli

“…Among the members of [NiFe]hydrogenases, O 2 -tolerant enzymes are particularly attractive as they remain catalytically active under oxic conditions, unlike all other hydrogenases [2]. This facilitates their biotechnological application in the areas of biohydrogen production, cofactor regeneration, and fuel cells [3][4][5]. All so-far isolated [NiFe]-hydrogenases consist of at least two subunits; a large subunit (LSU) of approximately 60 kDa housing the catalytic center and a small subunit (SSU) of approximately 30 kDa hosting one to three iron-sulfur (Fe-S) clusters [6].…”

Investigation of the Ferredoxin’s Influence on the Anaerobic and Aerobic, Enzymatic H2 Production

“…Hydrogenases can receive electrons from reduced ferredoxins (Fd red ) and combine them with protons to produce H 2 . The enzymatic activity of hydrogenases, especially the [FeFe] subtype proficient in H 2 production such as the one from the C. pasteurianum (CpI), can be exploited for biological H 2 production (Lu and Koo, 2019); wide deployment of the technology can contribute to reduction of CO 2 emission while supplying H 2 . Toward this aim, researchers have successfully developed both fermentative and photosynthetic pathways for reducing ferredoxins, which can subsequently be used for the enzymatic H 2 production (Smith et al, 2011;Yacoby et al, 2011).…”

“…Since Fd red is the source of electrons for the enzymatic H 2 production, presence of O 2 results in a lower H 2 yield via decreasing the amount of electrons delivered to the hydrogenases (Benemann et al, 1973;Koo and Swartz, 2018). O 2 also drives inactivation of the hydrogenases, which is irreversible and fast (within minutes under the atmospheric [O 2 ]) for most [FeFe] kinds (Lu and Koo, 2019). By fusing ferredoxin to an [FeFe] hydrogenase, Eilenberg et al (2016) successfully increased the photosynthetic H 2 production in the presence of O 2 , proposing that Fd red delayed inactivation of the hydrogenase at the expense of electrons.…”

Monitoring the Physiological State in the Dark Fermentation of Maize/Grass Silage Using Flow Cytometry and Electrooptic Polarizability Measurements

“…The photofermentation and photobiological processes require very specific cultivation conditions and have low efficiencies of light conversion and relatively low H 2 yields [2,13]. Enzymatic systems suffer from sensitivity to O 2 [8,14] and H 2 , which reduces the efficiency of these systems, although novel approaches are actively being researched to overcome this drawback [15]. Pure fermentative cultures (natural or engineered strains) require high sterilization costs that are currently unsustainable [9] and can lead to a negative net energy balance [8].…”