Escherichia coli can hardly grow anaerobically on glycerol without exogenous electron acceptor. The formate-consuming methanogen Methanobacterium formicicum plays a role as a living electron acceptor in glycerol fermentation of E. coli. Wild-type and mutant E. coli strains were screened for succinate production using glycerol in a co-culture with M. formicicum. Subsequently, E. coli was adapted to glycerol fermentation over 39 rounds (273 days) by successive co-culture with M. formicicum. The adapted E. coli (19.9 mM) produced twice as much succinate as non-adapted E. coli (9.7 mM) and 62% more methane. This study demonstrated improved succinate production from waste glycerol using an adapted wild-type strain of E. coli with wild-type M. formicicum, which is more useful than genetically modified strains. Crude glycerol, an economical feedstock, was used for the cultivation. Furthermore, the increase in methane production by M. formicicum during co-culture with adapted E. coli illustrated the possibility of energy-saving effects for the fermentation process.Electronic supplementary materialThe online version of this article (10.1007/s10295-017-1994-0) contains supplementary material, which is available to authorized users.

The production of gamma-aminobutyric acid (GABA) using GABA-producing lactic acid bacteria (LAB) has been considered to be an attractive strategy. However, some LAB may produce biogenic amines (BA), which may be of concern from the safety viewpoint. The aim of the present study was to characterize the production of GABA and BA in the soybean pastes fermented by Aspergillus oryzae (A. oryzae) FMB S46471 and GABA-producing Lactobacillus brevis (L. brevis) GABA 100. After a ripening period of 90 days, the levels of BA (putrescine, cadaverine, histamine, and tyramine) and GABA in the fermented soybean were assessed by highperformance liquid chromatography. The soybean pastes fermented by A. oryzae and L. brevis showed a range of 7,130-11,592 mg/kg for GABA, 178-305 mg/kg for tyramine, 139-163 mg/kg for putrescine, 7.4-10.8 mg/kg for histamine, and 7.1-7.9 mg/kg for cadaverine, whereas the soybean pastes fermented by A. oryzae only showed a range of 30-1,671 mg/kg for GABA, 0.8-189 mg/kg for tyramine, 1.3-85 mg/kg for putrescine, up to 3.6 mg/kg for histamine, and 0.2-2.4 mg/kg for cadaverine. The results showed that the production of GABA was accompanied by the increase in the production of BA, even though the production levels of histamine and cadaverine were very low. This is the first study to simultaneously characterize the production of BA and GABA in GABA-enriched fermented soybean pastes, and warrants further study to minimize the production of BA while optimizing the production of GABA.

Here, we demonstrate the ability of E. coli to perform glycerol fermentation in coculture with the methanogen M. formicicum to produce succinate. We found that the production of succinate by E. coli significantly increased during successive cocultivation.

We developed a formate-removing methanogenic inoculum (FRI) to facilitate succinate production from crude glycerol by Escherichia coli. FRI converted formate to methane, thereby enabling glycerol fermentation without additional electron acceptors under neutral pH. FRI was selectively enriched from sludge from the anaerobic digester of the Seonam sewage treatment plant (Seoul); this process was assessed via Illumina sequencing and scanning electron microscopy imaging. Methanobacterium congolense species occupied only 0.3% of the archaea community in the sludge and was enriched to 99.5% in complete FRI, wherein succinate-degrading bacteria were successfully eliminated. Co-culture with FRI improved glycerol fermentation and yielded 7.3 mM succinate from 28.7 mM crude glycerol, whereby FRI completely converted formate into methane. This study is the first to demonstrate methane production by M. congolense species, using formate. M. congolense-dominated FRI can serve as a renewable facilitator of waste feedstock fermentation and enable the production of commercially important compounds.

Metabolons represent the structural organization of proteins for metabolic or regulatory pathways. Here the interaction of fumarase FumB, aspartase AspA, and L-tartrate dehydratase TtdAB with the C4-dicarboxylate (C4-DC) transporters DcuA, DcuB, DcuC, and the L-tartrate transporter TtdT of Escherichia coli was tested by a bacterial two-hybrid (BACTH) assay in situ, or by co-chromatography using mSPINE (membrane Streptavidin protein interaction experiment). From the general C4-DC transporters, DcuB interacted with FumB and AspA, DcuA with AspA, whereas DcuC interacted with neither FumB nor AspA. Moreover, TtdT did not interact with TtdAB. The fumB-dcuB, the dcuA-aspA, and the ttdAB-ttdT genes encoding the respective proteins co-localize on the genome and each pair of genes forms co-transcripts whereas the dcuC gene lies alone. The data suggest the formation of DcuB/FumB and DcuB/AspA metabolons for the uptake of L-malate, or L-aspartate, and their conversion to fumarate for fumarate respiration and excretion of the product succinate. The DcuA/AspA metabolon catalyzes uptake and conversion of L-Asp to fumarate coupled to succinate excretion. The DcuA/AspA metabolon provides ammonia at the same time for nitrogen assimilation (ammonia shuttle). On the other hand, TtdT and TtdAB are not organized in a metabolon. Reasons for the formation (DcuA/AspA, DcuB/FumB, DcuB/AspA) or non-formation (DcuC, TtdT and TtdAB) of metabolons are discussed based on their metabolic roles.