Distinct Energy-Coupling Factor Transporter Subunits Enable Flavin Acquisition and Extracytosolic Trafficking for Extracellular Electron Transfer in Listeria monocytogenes

Rivera‐Lugo, Rafael; Huang, Shuo; Lee, Frank; Méheust, Raphaël; Iavarone, Anthony T.; Sidebottom, Ashley M.; Oldfield, Eric; Portnoy, Daniel A.; Light, S.H.

doi:10.1128/mbio.03085-22

Cited by 10 publications

(3 citation statements)

References 58 publications

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“…We performed comparative genomic analyses mining apbE gene clusters that lack a known electron transfer mechanism and found that a transmembrane protein with a DUF4405 domain of unknown function frequently colocalized with apbE in Proteobacteria and Firmicutes species ( Figure 5B ). Consistent with these DUF4405s functioning in flavinylation-based electron transfer, we observed gene clusters containing apbE and DUF4405 genes also often encoded genes for a flavinylated Flavodoxin and a transporter that provisions Listeria monocytogenes with extracytosolic flavins ( Figure 5C ) (Rivera-Lugo et al 2023).…”

Section: Resultsmentioning

confidence: 53%

Versatile roles of protein flavinylation in bacterial extracyotosolic electron transfer

Huang,

Méheust,

Barquera

et al. 2024

Preprint

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Bacteria perform diverse redox chemistries in the periplasm, cell wall, and extracellular space. Electron transfer for these extracytosolic activities is frequently mediated by proteins with covalently bound flavins, which are attached through post-translational flavinylation by the enzyme ApbE. Despite the significance of protein flavinylation to bacterial physiology, the basis and function of this modification remains unresolved. Here we apply genomic context analyses, computational structural biology, and biochemical studies to address the role of ApbE flavinylation throughout bacterial life. We find that ApbE flavinylation sites exhibit substantial structural heterogeneity. We identify two novel classes of flavinylation substrates that are related to characterized proteins with non-covalently bound flavins, providing evidence that protein flavinylation can evolve from a non-covalent flavoprotein precursor. We further find a group of structurally related flavinylation-associated cytochromes, including those with the domain of unknown function DUF4405, that presumably mediate electron transfer in the cytoplasmic membrane. DUF4405 homologs are widespread in bacteria and related to ferrosome iron storage organelle proteins that may facilitate iron redox cycling within ferrosomes. These studies reveal a complex basis for flavinylated electron transfer and highlight the discovery power of coupling comparative genomic analyses with high-quality structural models.

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Section: Resultsmentioning

confidence: 53%

Versatile roles of protein flavinylation in bacterial extracyotosolic electron transfer

Huang,

Méheust,

Barquera

et al. 2024

Preprint

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“…This is probably because there are many variations in EET gene configuration among members of the phylum Bacillota. Variations in the configuration of EET genes have been reported [16,21,22].…”

Section: Discussionmentioning

confidence: 99%

Effect of Fermentation Scale on Microbiota Dynamics and Metabolic Functions for Indigo Reduction

Farjana,

Furukawa,

Sumi

et al. 2023

IJMS

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During indigo dyeing fermentation, indigo reduction for the solubilization of indigo particles occurs through the action of microbiota under anaerobic alkaline conditions. The original microbiota in the raw material (sukumo: composted indigo plant) should be appropriately converged toward the extracellular electron transfer (EET)-occurring microbiota by adjusting environmental factors for indigo reduction. The convergence mechanisms of microbiota, microbial physiological basis for indigo reduction, and microbiota led by different velocities in the decrease in redox potential (ORP) at different fermentation scales were analyzed. A rapid ORP decrease was realized in the big batch, excluding Actinomycetota effectively and dominating Alkalibacterium, which largely contributed to the effective indigo reduction. Functional analyses of the microbiota related to strong indigo reduction on approximately day 30 indicated that the carbohydrate metabolism, prokaryotic defense system, and gene regulatory functions are important. Because the major constituent in the big batch was Alkalibacterium pelagium, we attempted to identify genes related to EET in its genome. Each set of genes for flavin adenine dinucleotide (FAD) transportation to modify the flavin mononucleotide (FMN)-associated family, electron transfer from NADH to the FMN-associated family, and demethylmenaquinone (DMK) synthesis were identified in the genome sequence. The correlation between indigo intensity reduction and metabolic functions suggests that V/A-type H+/Na+-transporting ATPase and NAD(P)H-producing enzymes drive membrane transportations and energization in the EET system, respectively.

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“…83 Recombinant reductase production and purification Hexa-histidine-tagged reductase constructs were expressed in E. coli Rosetta cells using either the pET28a or pMCSG53 expression vectors, as previously described. 84 Briefly, expression was performed using 500 mL cultures of Luria-Bertani broth (BD LB Broth, Miller #244610) + 100 µM riboflavin, grown to an OD of 0.7-1.0 at 37 °C, then induced with 1 mM β-d-1-thiogalactopyranoside overnight at 20 °C. Kanamycin (30 mg/mL, pET28a vector) or carbenicillin (100 mg/mL, pMCSG53 vector) were used for selection.…”

Section: Proteomics Mass Spectrometrymentioning

confidence: 99%

Dietary- and host-derived metabolites are used by diverse gut bacteria for anaerobic respiration

Little

Younker

Schechter

et al. 2022

Preprint

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Anaerobic respiration encompasses a major class of microbial energy metabolisms that employ reductases to respire different non-oxygen electron acceptors. Respiratory reductases play important roles in multiple geochemical cycles but their significance in other contexts remains unclear. Here we identify three taxonomically distinct families of gut bacteria that encode exceptionally large arsenals of tens to hundreds of respiratory-like reductases per genome. By screening representative species from each family, we discover 22 metabolites used as respiratory electron acceptors in a species-specific manner. Identified respiratory reactions transform multiple classes of dietary- and host-derived fecal metabolites, including bioactive molecules like resveratrol and the immunometabolite itaconate. Reductase substrate-profiling identifies enzyme-substrate pairs and paints a striking picture of reductase evolution, including evidence that reductases with specificities for related cinnamate substrates but distinct mechanisms independently emerged at least four separate times. These studies define an exceptionally versatile form of respiration that directly links microbial central metabolism to the gut metabolome.

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Distinct Energy-Coupling Factor Transporter Subunits Enable Flavin Acquisition and Extracytosolic Trafficking for Extracellular Electron Transfer in Listeria monocytogenes

Cited by 10 publications

References 58 publications

Versatile roles of protein flavinylation in bacterial extracyotosolic electron transfer

Versatile roles of protein flavinylation in bacterial extracyotosolic electron transfer

Effect of Fermentation Scale on Microbiota Dynamics and Metabolic Functions for Indigo Reduction

Dietary- and host-derived metabolites are used by diverse gut bacteria for anaerobic respiration

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