Electrochemical Studies of CO<sub>2</sub>‐Reducing Metalloenzymes

Meneghello, Marta; Léger, Christophe; Fourmond, Vincent

doi:10.1002/chem.202102702

Cited by 14 publications

(16 citation statements)

References 121 publications

(165 reference statements)

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“…(2) All metallo-FDHs are thought to be capable of catalyzing the CO 2 reduction reaction (CO2RR) . However, there is no known biocatalyst that accomplishes CO2RR in air . Also, it is not possible to test this reaction with low-potential artificial electron donors (e.g., MV) because they react with O 2 .…”

Section: Resultsmentioning

confidence: 99%

How a Formate Dehydrogenase Responds to Oxygen: Unexpected O₂ Insensitivity of an Enzyme Harboring Tungstopterin, Selenocysteine, and [4Fe–4S] Clusters

et al. 2022

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The reversible two-electron interconversion of formate and CO2 is catalyzed by both nonmetallo- and metallo-formate dehydrogenases (FDHs). The latter group comprises molybdenum- or tungsten-containing enzymes with the metal coordinated by two equivalents of a pyranopterin cofactor, a cysteinyl or selenocysteinyl (Sec) ligand supplied by the polypeptide, and a catalytically essential terminal sulfido ligand. In addition, these biocatalysts incorporate one or more [4Fe–4S] clusters for facilitating long-distance electron transfer. However, an interesting dichotomy arises when attempting to understand how the metallo-FDHs react with O2. Whereas existing scholarship portrays these enzymes as being unable to perform in air due to extreme O2 lability of their metal centers, studies dating as far back as the 1930s emphasize that some of these systems exhibit formate oxidase (FOX) activity, coupling formate oxidation to O2 reduction. Therefore, to reconcile these conflicting views, we explored context-dependent functional linkages between metallo-FDHs and their cognate electron acceptors within the same organism vis-à-vis catalysis under atmospheric O2. Here, we report the discovery and characterization of an O2-insensitive FDH2 from the sulfate-reducing bacterium Desulfovibrio vulgaris Hildenborough (DvH) that ligates tungsten, Sec, and four [4Fe–4S] clusters. By advancing a robust expression platform for its recombinant production, we eliminate both the requirement of nitrate or azide during purification and reductive activation with thiols and/or formate prior to catalysis. Because the distinctive spectral signatures of formate-reduced DvH-FDH2 remain invariant under anaerobic and aerobic conditions, we benchmarked the enzyme activity in air, identifying CO2 as the catalytic product. Full reaction progress curve analysis discloses a high catalytic efficiency when probed with a high-potential artificial electron acceptor. Furthermore, we show that DvH-FDH2 enables near-stoichiometric hydrogen peroxide production without superoxide release to achieve O2 insensitivity. Notably, simultaneous electron transfer to cytochrome c and O2 reveals that metal-based electron bifurcation is operational in this system. Taken together, our work proves the co-occurrence of redox bifurcated FDH and FOX activities within a metalloenzyme scaffold. These findings set the stage for uncovering previously unknown O2-insensitive flavin-based electron bifurcation mechanisms, as well as for developing authentic formate/air biofuel cells, engineering O2-stable FDHs and biohybrid metallocatalysts, and discerning formate bioenergetics of gut microbiota.

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Section: Resultsmentioning

confidence: 99%

How a Formate Dehydrogenase Responds to Oxygen: Unexpected O₂ Insensitivity of an Enzyme Harboring Tungstopterin, Selenocysteine, and [4Fe–4S] Clusters

et al. 2022

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“…Additionally, the analysis of “catalytic” currents can be followed via protein film electrochemistry (Table ). , …”

Section: Formate Dehydrogenasementioning

confidence: 99%

Second and Outer Coordination Sphere Effects in Nitrogenase, Hydrogenase, Formate Dehydrogenase, and CO Dehydrogenase

et al. 2022

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Gases like H2, N2, CO2, and CO are increasingly recognized as critical feedstock in “green” energy conversion and as sources of nitrogen and carbon for the agricultural and chemical sectors. However, the industrial transformation of N2, CO2, and CO and the production of H2 require significant energy input, which renders processes like steam reforming and the Haber-Bosch reaction economically and environmentally unviable. Nature, on the other hand, performs similar tasks efficiently at ambient temperature and pressure, exploiting gas-processing metalloenzymes (GPMs) that bind low-valent metal cofactors based on iron, nickel, molybdenum, tungsten, and sulfur. Such systems are studied to understand the biocatalytic principles of gas conversion including N2 fixation by nitrogenase and H2 production by hydrogenase as well as CO2 and CO conversion by formate dehydrogenase, carbon monoxide dehydrogenase, and nitrogenase. In this review, we emphasize the importance of the cofactor/protein interface, discussing how second and outer coordination sphere effects determine, modulate, and optimize the catalytic activity of GPMs. These may comprise ionic interactions in the second coordination sphere that shape the electron density distribution across the cofactor, hydrogen bonding changes, and allosteric effects. In the outer coordination sphere, proton transfer and electron transfer are discussed, alongside the role of hydrophobic substrate channels and protein structural changes. Combining the information gained from structural biology, enzyme kinetics, and various spectroscopic techniques, we aim toward a comprehensive understanding of catalysis beyond the first coordination sphere.

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“…[9b] The direct reduction of CO 2 is particularly noteworthy, as it places nitrogenases in a small group of enzymes with the same ability, i. e. nickel-iron CO dehydrogenases and formate (HCOO À ) dehydrogenases, which catalyze the reversible reduction of CO 2 to CO and HCOO À , respectively. [40] Beyond the biological importance of these reactivities, the conversion of CO 2 into hydrocarbons suggests an industrial use of nitrogenases for carbon capture and the production of biofuels in a green economy. Due to the increasing importance of carbon capture and biofuel production, we review and discuss the current knowledge of the CO and CO 2 conversion by all three nitrogenase isozymes: the Mo, V and Fe-only nitrogenase.…”

Section: Substrate Range Of Nitrogenasesmentioning

confidence: 99%

“…Furthermore, the reduction of CO 2 to methane (CH 4 ) by nitrogenases has spiked interest for its potential role in shaping microbial communities [9b] . The direct reduction of CO 2 is particularly noteworthy, as it places nitrogenases in a small group of enzymes with the same ability, i. e. nickel‐iron CO dehydrogenases and formate (HCOO − ) dehydrogenases, which catalyze the reversible reduction of CO 2 to CO and HCOO − , respectively [40] …”

Section: Introductionmentioning

confidence: 99%

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The Conversion of Carbon Monoxide and Carbon Dioxide by Nitrogenases

Oehlmann

Rebelein

2021

ChemBioChem

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Nitrogenases are the only known family of enzymes that catalyze the reduction of molecular nitrogen (N2) to ammonia (NH3). The N2 reduction drives biological nitrogen fixation and the global nitrogen cycle. Besides the conversion of N2, nitrogenases catalyze a whole range of other reductions, including the reduction of the small gaseous substrates carbon monoxide (CO) and carbon dioxide (CO2) to hydrocarbons. However, it remains an open question whether these ‘side reactivities’ play a role under environmental conditions. Nonetheless, these reactivities and particularly the formation of hydrocarbons have spurred the interest in nitrogenases for biotechnological applications. There are three different isozymes of nitrogenase: the molybdenum and the alternative vanadium and iron‐only nitrogenase. The isozymes differ in their metal content, structure, and substrate‐dependent activity, despite their homology. This minireview focuses on the conversion of CO and CO2 to methane and higher hydrocarbons and aims to specify the differences in activity between the three nitrogenase isozymes.

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Electrochemical Studies of CO₂‐Reducing Metalloenzymes

Cited by 14 publications

References 121 publications

How a Formate Dehydrogenase Responds to Oxygen: Unexpected O₂ Insensitivity of an Enzyme Harboring Tungstopterin, Selenocysteine, and [4Fe–4S] Clusters

How a Formate Dehydrogenase Responds to Oxygen: Unexpected O₂ Insensitivity of an Enzyme Harboring Tungstopterin, Selenocysteine, and [4Fe–4S] Clusters

Second and Outer Coordination Sphere Effects in Nitrogenase, Hydrogenase, Formate Dehydrogenase, and CO Dehydrogenase

The Conversion of Carbon Monoxide and Carbon Dioxide by Nitrogenases

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Electrochemical Studies of CO2‐Reducing Metalloenzymes

Cited by 14 publications

References 121 publications

How a Formate Dehydrogenase Responds to Oxygen: Unexpected O2 Insensitivity of an Enzyme Harboring Tungstopterin, Selenocysteine, and [4Fe–4S] Clusters

How a Formate Dehydrogenase Responds to Oxygen: Unexpected O2 Insensitivity of an Enzyme Harboring Tungstopterin, Selenocysteine, and [4Fe–4S] Clusters

Second and Outer Coordination Sphere Effects in Nitrogenase, Hydrogenase, Formate Dehydrogenase, and CO Dehydrogenase

The Conversion of Carbon Monoxide and Carbon Dioxide by Nitrogenases

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Product

Resources

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Electrochemical Studies of CO₂‐Reducing Metalloenzymes

How a Formate Dehydrogenase Responds to Oxygen: Unexpected O₂ Insensitivity of an Enzyme Harboring Tungstopterin, Selenocysteine, and [4Fe–4S] Clusters

How a Formate Dehydrogenase Responds to Oxygen: Unexpected O₂ Insensitivity of an Enzyme Harboring Tungstopterin, Selenocysteine, and [4Fe–4S] Clusters