The modified Q-cycle: A look back at its development and forward to a functional model

Crofts, Antony R.

doi:10.1016/j.bbabio.2021.148417

Cited by 30 publications

(30 citation statements)

References 210 publications

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“…Some have used the term cooperative bidirectional one-by-one electron transfer to distinguish it from noncooperative counterparts, as well as unidirectional events . EB was originally proposed in 1976 to rationalize mitochondrial quinone-based (QB) energy coupling. , The present-day literature is largely focused on flavin-based (FB) EB, − which is believed to have been discovered in 2008. However, one of us (C.S.R.)…”

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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“…aa 3 or complex IV, CIV). After exchange of Q for QH 2 at the Q P site of CIII, the sequence of events is repeated, resulting in formation of QH 2 at the Q N site and abstraction of two protons from the n side of the membrane, contributing further to the PMF (for review, see Berry et al, 2013;Brzezinski et al, 2021;Cramer et al, 2011;Crofts, 2004;Mulkidjanian, 2005;Sarewicz and Osyczka, 2015;Sarewicz et al, 2021;Crofts, 2021).…”

Section: Introductionmentioning

confidence: 99%

The respiratory supercomplex from C. glutamicum

et al. 2022

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“…The structural characterisation at high resolution sheds light on the mechanism of the cyt bcc - aa 3 supercomplex. In contrast to all other bc complexes known so far, which enable bifurcated electron transfer with a mobile Rieske protein 1 , 52 , the homologous subunit QcrA is integrated into the supercomplex as a non-mobile subunit (Fig. 4a, b ).…”

Section: Discussionmentioning

confidence: 92%

“…4a, b ). Yet, the cyt bcc complex fulfils the prerequisite for a Q cycle 1 , 52 having a Q o and a Q i site, which were mapped by the transition state analogue stigmatellin and a natively co-purified menaquinone, respectively (Fig. 2b ).…”

Section: Discussionmentioning

confidence: 99%

Structural basis for safe and efficient energy conversion in a respiratory supercomplex

et al. 2022

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Proton-translocating respiratory complexes assemble into supercomplexes that are proposed to increase the efficiency of energy conversion and limit the production of harmful reactive oxygen species during aerobic cellular respiration. Cytochrome bc complexes and cytochrome aa3 oxidases are major drivers of the proton motive force that fuels ATP generation via respiration, but how wasteful electron- and proton transfer is controlled to enhance safety and efficiency in the context of supercomplexes is not known. Here, we address this question with the 2.8 Å resolution cryo-EM structure of the cytochrome bcc-aa3 (III2-IV2) supercomplex from the actinobacterium Corynebacterium glutamicum. Menaquinone, substrate mimics, lycopene, an unexpected Qc site, dioxygen, proton transfer routes, and conformational states of key protonable residues are resolved. Our results show how safe and efficient energy conversion is achieved in a respiratory supercomplex through controlled electron and proton transfer. The structure may guide the rational design of drugs against actinobacteria that cause diphtheria and tuberculosis.

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The modified Q-cycle: A look back at its development and forward to a functional model

Cited by 30 publications

References 210 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

The respiratory supercomplex from C. glutamicum

Structural basis for safe and efficient energy conversion in a respiratory supercomplex

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The modified Q-cycle: A look back at its development and forward to a functional model

Cited by 30 publications

References 210 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

The respiratory supercomplex from C. glutamicum

Structural basis for safe and efficient energy conversion in a respiratory supercomplex

Contact Info

Product

Resources

About

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