Ferric iron and extracellular electron shuttling increase xylose utilization and butanol production during fermentation with multiple solventogenic bacteria

Abstract

Xylose is the second most abundant sugar derived from lignocellulose; it is considered less desirable than glucose for fermentation, and strategies that specifically increase xylose utilization in wild-type cells are goals for biofuel production. Xylose consumption, butanol production, and hydrogen production increased in both Clostridium beijerinckii and a novel solventogenic bacterium (strain DC-1) when anthraquinone-2,6,-disulfonate (AQDS) or riboflavin were used as redox mediators to transfer electrons to poorly crystalline Fe(OH) as an extracellular electron sink. Strain DC-1 was most closely related to Rhizobiales bacterium Mfc52 based on 95% 16S rRNA gene sequence similarity, which demonstrates that this response is not limited to a single genus of xylose-fermenting bacteria. Xylose utilization and butanol production were negligible in control incubations containing cells plus 3% (w/v) xylose alone during a 10-day batch fermentation, for both strains tested (n-butanol titers of 0.05 g L). Micromolar concentrations of AQDS and riboflavin were added as electron shuttling compounds with poorly crystalline Fe(OH) as an insoluble electron acceptor, and respective n-butanol titers increased to 6.35 and 7.46 g L. Increases in xylose consumption for the iron treatments were relatively high, from less than 0.49 g L (xylose alone, no iron or electron shuttling molecules) to 25.98 and 29.15 g L for the AQDS and riboflavin treatments, respectively. Hydrogen production was also 3.68 times greater for the AQDS treatment and 5.27 greater for the riboflavin treatment relative to controls. Strain DC-1 data were similar, again indicating that the effects are not specific to the genus Clostridium.

Overview on the Bacterial Iron-Riboflavin Metabolic Axis

“…Geothrix fermentans , an isolate of an aquifer contaminated with petroleum, uses secreted riboflavin to shuttle electrons toward environmental Fe 3+ ( Mehta-Kolte and Bond, 2012 ). Carbohydrate oxidation is increased when using riboflavin as electron transfer mediator in the presence of crystalline Fe(OH) 3 as extracellular electron sink in Clostridium beijerinckii and an uncharacterized novel rhizobial solventogenic bacterium ( Popovic et al, 2017 ). Also, riboflavin is likely involved in the reduction of extracellular Fe 3+ -citrate and solid phase hydrous ferric oxide by Desulfotomaculum reducens , a sulfate-reducing Gram-positive species ( Dalla Vecchia et al, 2014 ) and in anaerobic Fe 3+ reduction by an alkaliphilic bacterial consortium ( Fuller et al, 2014 ).…”

Enhancement of Butanol Production: From Biocatalysis to Bioelectrocatalysis

“…Specifically, one is to improve redox metabolism that is highly associated with the flow of reducing power required for biosynthesis of butanol, which is mainly 15,16 The other is to facilitate ET for enhanced bioelectrocatalysis through the integration of the optimized BES that can directly supply the bioelectrocatalysis with electrons on the electrodes and/or the introduction of exogenous redox mediators (both natural and synthetic) that can improve ET and/or redox balance for an enhanced bioconversion of butanol in bioreactors. 7,17,18 Finally, we give some own remarks and perspectives on this field for future improvement.…”

“…56,71,72 Furthermore, an increase in both xylose utilization and butanol production was obtained in C. beijerinckii by using anthraquinone-2,6,-disulfonate (AQDS) or riboflavin as redox mediators to facilitate extracellular electron transfer. 17 On the other hand, many studies have also reported that the supplement of redox mediators in fermentation media could enhance butanol production by redirecting metabolic flux to butanol synthesis or by balancing intracellular redox metabolism through improving the availability of NADH (Figure 6A). Du et al demonstrated that supplementing 500 mM MV decreased hydrogen, acetate, and butyrate production by more than 80− 90%, while it increased butanol production by more than 40% in a mutant C. tyrobutyricum Δack-adhE2 (Figure 6B,C).…”

“…Then, we expand the discussion by targeting the two limiting factors: redox balance and ET. Specifically, one is to improve redox metabolism that is highly associated with the flow of reducing power required for biosynthesis of butanol, which is mainly mediated by diffusional cofactors (i.e., NAD + /NADH and NADP + /NADPH) and specific oxidoreductases. , The other is to facilitate ET for enhanced bioelectrocatalysis through the integration of the optimized BES that can directly supply the bioelectrocatalysis with electrons on the electrodes and/or the introduction of exogenous redox mediators (both natural and synthetic) that can improve ET and/or redox balance for an enhanced bioconversion of butanol in bioreactors. ,, Finally, we give some own remarks and perspectives on this field for future improvement. Enhancement of butanol bioconversion should begin with a review of the nature of biochemical processes.…”

“…The addition of such electron carriers could divert electrons from ferredoxin used for hydrogen formation to NADH accumulation by affecting the activities of hydrogenase and NAD + reductase, resulting in redox balance and an increased carbon flux toward butanol synthesis (Figure A). ,, Furthermore, an increase in both xylose utilization and butanol production was obtained in C. beijerinckii by using anthraquinone-2,6,-disulfonate (AQDS) or riboflavin as redox mediators to facilitate extracellular electron transfer . On the other hand, many studies have also reported that the supplement of redox mediators in fermentation media could enhance butanol production by redirecting metabolic flux to butanol synthesis or by balancing intracellular redox metabolism through improving the availability of NADH (Figure A).…”