Flavodoxin hydroquinone provides electrons for the ATP‐dependent reactivation of protein‐bound corrinoid cofactors

Abstract

Corrinoid-dependent enzyme systems rely on the super-reduced state of the protein-bound corrinoid cofactor to be functional, for example, in methyl transfer reactions. Due to the low redox potential of the [Co II ]/[Co I ] couple, autoxidation of the corrinoid cofactor occurs and leads to the formation of the inactive [Co II ]-state. For the reactivation, which is an energydemanding process, electrons have to be transferred from a physiological donor to the corrinoid cofactor by the help of a reductive activator protein. In this study, we identified reduced flavodoxin as electron donor for the ATP-dependent reduction of protein-bound corrinoid cofactors of bacterial O-demethylase enzyme systems. Reduced flavodoxin was generated enzymatically using pyruvate:ferredoxin/flavodoxin oxidoreductase rather than hydrogenase. Two of the four flavodoxins identified in Acetobacterium dehalogenans and Desulfitobacterium hafniense DCB-2 were functional in supplying electrons for corrinoid reduction. They exhibited a midpoint potential of about À400 mV (E SHE , pH 7.5) for the semiquinone/ hydroquinone transition. Reduced flavodoxin could be replaced by reduced clostridial ferredoxin. It was shown that the low-potential electrons of reduced flavodoxin are first transferred to the iron-sulfur cluster of the reductive activator and finally to the protein-bound corrinoid cofactor. This study further highlights the importance of reduced flavodoxin, which allows maintaining a variety of enzymatic reaction cycles by delivering low-potential electrons.

Biochemistry of methanol‐dependent acetogenesis in Eubacterium callanderi KIST612

“…Since corrinoids are highly reactive in their superreduced active state, they undergo spontaneous autooxidation leading to an inactivation of the CoP. To return the corrinoid cofactor to the active state, so-called activating enzymes use ferredoxin/ flavodoxin to reductively activate the corrinoid protein (Dürichen et al, 2019;Kißling et al, 2020). MTI confers the substrate specificity and a BLAST search using the Eubacterium limosum MTI (MttB, WP_038351887.1; (Kountz et al, 2020)) as query showed that the genome of E. callanderi KIST612 encodes 32 different MTI enzymes (Table S1), underlining the importance of methyl groups as carbon and energy source for E. callanderi KIST612.…”

Gut Metabolism of Furanocoumarins: Proposed Function of Co O-Methyltransferase

“…The unique methyl ether cleavage is achieved by the action of supernucleophilic Co(I) reactive species generated in the Co-corrinoid O-methyltrans- ferase enzyme. 24 Since three A. dahurica furanocoumarins, compounds 1, 2, and 6 in Figure 1, contain methyl aryl ether functional group, each furanocoumarin was reacted to Blautia sp. MRG-PMF1 strain to examine whether the strain is responsible for the conversion.…”

“…MRG-PMF1 strain was reported to metabolize polymethoxyflavones to the corresponding flavones by the cleavage of methyl aryl ether groups. The unique methyl ether cleavage is achieved by the action of supernucleophilic Co­(I) reactive species generated in the Co-corrinoid O -methyltransferase enzyme . Since three A.…”

Sustainable B12-Dependent Dehalogenation of Organohalides in E. coli

“…Encouraged by reductive dehalogenation in the presence of added riboflavin, we tested more flavin mononucleotide (FMN)-binding flavodoxins (Flds) for suitability as electron shuttlers. We expressed and purified five flavodoxins from E. coli (EcFld), 43 Azobacter vinelandii (AvFld), 44,45 Chlorobium phaeovibroides (CpFld), 46 Acetobacterium dehalogenans (AdFld), 47 , and D. hafniense DCB-2 (DhFld). 47 Purified Flds were largely apo and were supplemented with FMN in biochemical assays.…”

“…We expressed and purified five flavodoxins from E. coli (EcFld), 43 Azobacter vinelandii (AvFld), 44,45 Chlorobium phaeovibroides (CpFld), 46 Acetobacterium dehalogenans (AdFld), 47 , and D. hafniense DCB-2 (DhFld). 47 Purified Flds were largely apo and were supplemented with FMN in biochemical assays. Formation of phenol was observed after incubating 50 μM DhFld with lysates containing DaPFOR and RdhC1 in the presence of necessary cofactors.…”