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  • Introduction Formate dehydrogenase enzymes FDHs are

    2020-09-18

    Introduction Formate dehydrogenase enzymes (FDHs) are a group of heterogeneous proteins that catalyse the reversible formate () oxidation to carbon dioxide () (Eq. (1)). These enzymes are classified in two families, one gathering the enzymes that hold no redox-active centres and another that includes all the metal-dependent enzymes [[1], [2], [3]]. The metal-dependent FDHs are found only in prokaryotic organisms and all have in common the presence of one molybdenum or one tungsten ion in the active site coordinated by four sulfur atoms of two pyranopterin cofactors and additional sulfur and/or selenium atoms [[4], [5], [6]]. Both FDH acetylcholine receptor have been the subject of intense research, since it was unveiled that the enzymatic reaction can be run reversibly, depending on the experimental conditions, allowing the consumption (reduction) of carbon dioxide with formation of formate (Eq. (1)) [[1], [2], [3],[7], [8], [9], [10], [11]]. This enzymatic reaction is of great interest since it may contribute to decrease the high atmospheric carbon dioxide levels, converting this green-house gas back into fuel or other added-value chemicals [12,13]. The reaction of formate formation from CO2 is one of the most interesting, because of the highest added value when compared with the energy required to form the product [14]. The reaction electrochemical control has been pursuit aiming an integration on bioelectrochemical systems (BES) that would allow the CO2 conversion with (sustainable) low energy consumption, employing mild conditions, to produce a single and specific product – formate (or formic acid) [15]. The feasibility of the utilization of FDHs, in general as catalyst in BES, has already been shown, namely using FDH isolated from Candida boidinii (an enzyme that is NADH-dependent and metal-independent, available commercially) in the electrosynthesis of formate from CO2, although the imposition of a high overpotential was necessary (−1 V vs. Ag/AgCl) [16]. A decrease in the required overpotential was described with the immobilization of Methylobacterium extorquens (Me) FDH (a tungsten-containing enzyme), through the use of modified carbon-based electrodes with several polymers incorporating mediators [17]. Recently, the co-immobilization of a redox polymer (cobaltocene) together with a molybdenum-containing FDH (isolated from Escherichia coli) allowed the enhancement of the three-dimension matrix and, as so, also the mediated electron transfer rate [18]. The use of direct electrochemical methods to tune the FDH activity has been widely searched [[19], [20], [21], [22]]. Amongst the studies of the viability in using FDH enzymes to catalyse the CO2 reduction, recently, it was found that the periplasmic FDH from Desulfovibrio desulfuricans () is one of the most efficient carbon dioxide reducers so far described [7]. The enzyme has been demonstrated to possess high affinity for CO2 (Kmapp = 15.7 μM) and high rates for its reduction (kcatapp = 46.6 s−1). DdFDH is a heterotrimeric enzyme, harbouring one molybdenum-containing active site, two [4Fe-4S] centres and four haems c. The molybdenum centre, in its oxidised form, harbours the molybdenum ion coordinated by the cis-dithiolene group of two pyranopterin molecules, one selenium from a selenium-cysteine residue and one terminal sulfur atom [23,24]. The other metallic redox-active centres, besides the Mo active site, have an electron transfer role, allowing the existence of an intramolecular electronic pathway from the physiological electron partner to the active site [25]. During catalysis, the molybdenum ion cycles between the Mo(VI) and Mo(IV) oxidation states and the formate oxidation/carbon dioxide reduction were recently suggested to proceed through hydride abstraction/insertion (respectively), with the terminal sulfido group of the molybdenum centre acting as the direct hydride acceptor (Mo(VI) = S) / donor (Mo(IV)-SH) (respectively) [[7], [8], [9], [10], [11]]. The two electrons (Eq. (1)) generated upon formate oxidation, or consumed during the carbon dioxide reduction are provided or accepted (respectively) by the physiological electron partner of each specific FDH enzyme. In vitro or in artificial systems, the electrons can be delivered/accepted by artificial electron donors/acceptors or by an electrode (as part of an electrochemical cell), via mediated or direct electron transfer [17,19,26]. Due to its unique kinetic parameters towards the carbon dioxide reduction [7], the DdFDH is a good candidate to be incorporated in direct bioelectrochemical systems. Aiming to develop a future BES, in this work, we have characterised the DdFDH electrochemical properties under non-turnover conditions, on pyrolytic graphite electrode, using direct electrochemical methods, without any mediators or surface modifiers. The electrochemical DdFDH catalytic response towards the carbon dioxide reduction was also observed. The use of direct gaseous CO2 injection into the electrochemical cell was attempted resulting in observable catalytic currents, showing to be a viable methodology.