Structure and function of an unusual flavodoxin from the domain Archaea

Abstract

Flavodoxins, electron transfer proteins essential for diverse metabolisms in microbes from the domainBacteria, are extensively characterized. Remarkably, although genomic annotations of flavodoxins are widespread in microbes from the domainArchaea, none have been isolated and characterized. Herein is described the structural, biochemical, and physiological characterization of an unusual flavodoxin (FldA) fromMethanosarcina acetivorans, an acetate-utilizing methane-producing microbe of the domainArchaea. In contrast to all flavodoxins, FldA is homodimeric, markedly less acidic, and stabilizes an anionic semiquinone. The crystal structure reveals an flavin mononucleotide (FMN) binding site unique from all other flavodoxins that provides a rationale for stabilization of the anionic semiquinone and a remarkably low reduction potentials for both the oxidized/semiquinone (−301 mV) and semiquinone/hydroquinone couples (−464 mV). FldA is up-regulated in acetate-grown versus methanol-grown cells and shown here to substitute for ferredoxin in mediating the transfer of low potential electrons from the carbonyl of acetate to the membrane-bound electron transport chain that generates ion gradients driving ATP synthesis. FldA offers potential advantages over ferredoxin by (i) sparing iron for abundant iron-sulfur proteins essential for acetotrophic growth and (ii) resilience to oxidative damage.

Methanosarcina acetivorans: A Model for Mechanistic Understanding of Aceticlastic and Reverse Methanogenesis

“…The acetyl-CoA is decarbonylated by the acetyl-CoA decarbonylase/synthase (ACDS) yielding a methyl group and CO. The methyl group is transferred to tetrahydrosarcinapterin (H 4 SPT) yielding CH 3 -H 4 SPT and CO is oxidized to CO 2 with transfer of electrons to either ferredoxin (Fdx) or a novel flavodoxin (FldA) characterized from Ms. acetivorans (Prakash et al, 2019b).…”

“…a potential advantage in periods of oxidative stress that damage the iron-sulfur clusters of Fdx, or when iron is limiting in the environment (Prakash et al, 2019b).…”

“…The F 420 H 2 dehydrogenase, essential for methylotrophic growth, is down regulated in acetate-grown cells leading to the proposal that oxidation of F 420 H 2 is dependent on the electron bifurcating HdrA 2 B 2 C 2 (Yan et al, 2017). As FldA can replace Fdx as electron acceptor for HdrA 2 B 2 C 2 , and donor to Rnf, either are available for initiating fermentative and respiratory electron transport (Prakash et al, 2019b). The combination of fermentative and respiratory electron transport generates both H + and Na + gradients that drive ATP synthesis by the ATP synthase dependent on both gradients (Schlegel et al, 2012a).…”

“…Several acetotrophic Methanosarcina lack Ech and Vho hydrogenases and are H 2 independent ( Zhilina, 1978 ; Ollivier et al, 1984 ; Sowers et al, 1984a ; Zinder et al, 1985 ; Elberson and Sowers, 1997 ; Von Klein et al, 2002 ; Shimizu et al, 2011 ; Ganzert et al, 2014 ). Ms. acetivorans is typical of H 2 independent Methanosarcina that instead utilize the membrane bound RnfCDGEAB complex to oxidize Fdx or FldA ( Figure 2B ; Li et al, 2006 ; Wang et al, 2011 ; Schlegel et al, 2012b ; Prakash et al, 2019b ). FldA accepts electrons from ACDS and is proposed to replace Fdx when growing in iron-limited environments ( Prakash et al, 2019b ).…”

“…Ms. acetivorans is typical of H 2 independent Methanosarcina that instead utilize the membrane bound RnfCDGEAB complex to oxidize Fdx or FldA ( Figure 2B ; Li et al, 2006 ; Wang et al, 2011 ; Schlegel et al, 2012b ; Prakash et al, 2019b ). FldA accepts electrons from ACDS and is proposed to replace Fdx when growing in iron-limited environments ( Prakash et al, 2019b ). Fdx is an electron donor to the RnfB subunit of the Rnf complex ( Suharti et al, 2014 ).…”