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A newly isolated Bacillus licheniformis strain thermophilically produces 2,3-butanediol, a platform and fuel bio-chemical.

ABSTRACT: BACKGROUND:2,3-Butanediol (2,3-BD), a platform and fuel bio-chemical, can be efficiently produced by Klebsiella pneumonia, K. oxytoca, and Serratia marcescens. However, these strains are opportunistic pathogens and not favorable for industrial application. Although some generally regarded as safe (GRAS) microorganisms have been isolated in recent years, there is still a demand for safe 2,3-BD producing strains with high productivity and yield under thermophilic fermentation. RESULTS:Bacillus licheniformis strain 10-1-A was newly isolated for 2,3-BD production. The optimum temperature and medium pH were 50°C and pH 7.0 for 2,3-BD production by strain 10-1-A. The medium composition was optimized through Plackett-Burman design and response surface methodology techniques. With a two-stage agitation speed control strategy, 115.7 g/L of 2,3-BD was obtained from glucose by fed-batch fermentation in a 5-L bioreactor with a high productivity (2.4 g/L·h) and yield (94% of its theoretical value). The 2,3-BD produced by strain 10-1-A comprises (2R,3R)-2,3-BD and meso-2,3-BD with a ratio of nearly 1:1. The bdh and gdh genes encoding meso-2,3-butanediol dehydrogenase (meso-BDH) and glycerol dehydrogenase (GDH) of strain 10-1-A were expressed in Escherichia coli and the proteins were purified. meso-2,3-BD and (2R,3R)-2,3-BD were transformed from racemic acetoin by meso-BDH and GDH with NADH, respectively. CONCLUSIONS:Compared with the reported GRAS 2,3-BD producers, B. licheniformis 10-1-A could thermophilically produce 2,3-BD with a high concentration, productivity and yield. Thus, the newly isolated GRAS strain 10-1-A might be a promising strain for industrial production of 2,3-BD. Two key enzymes for meso-2,3-BD and (2R,3R)-2,3-BD production were purified and further studied, and this might be helpful to understand the mechanism for 2,3-BD stereoisomers forming in B. licheniformis.

SUBMITTER: Li L

PROVIDER: S-EPMC3766113 | biostudies-literature | 2013 Aug

REPOSITORIES: biostudies-literature

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A newly isolated Bacillus licheniformis strain thermophilically produces 2,3-butanediol, a platform and fuel bio-chemical.

Li Lixiang L Zhang Lijie L Li Kun K Wang Yu Y Gao Chao C Han Binbin B Ma Cuiqing C Xu Ping P

Biotechnology for biofuels 20130828 1

<h4>Background</h4>2,3-Butanediol (2,3-BD), a platform and fuel bio-chemical, can be efficiently produced by Klebsiella pneumonia, K. oxytoca, and Serratia marcescens. However, these strains are opportunistic pathogens and not favorable for industrial application. Although some generally regarded as safe (GRAS) microorganisms have been isolated in recent years, there is still a demand for safe 2,3-BD producing strains with high productivity and yield under thermophilic fermentation.<h4>Results</ ...[more]

PMID: 23981315

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Project description:BackgroundPreviously, a safe strain, Bacillus amyloliquefaciens B10-127 was identified as an excellent candidate for industrial-scale microbial fermentation of 2,3-butanediol (2,3-BD). However, B. amyloliquefaciens fermentation yields large quantities of acetoin, lactate and succinate as by-products, and the 2,3-BD yield remains prohibitively low for commercial production.Methodology/principal findingsIn the 2,3-butanediol metabolic pathway, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) catalyzes the conversion of 3-phosphate glyceraldehyde to 1,3-bisphosphoglycerate, with concomitant reduction of NAD(+) to NADH. In the same pathway, 2,3-BD dehydrogenase (BDH) catalyzes the conversion of acetoin to 2,3-BD with concomitant oxidation of NADH to NAD(+). In this study, to improve 2,3-BD production, we first over-produced NAD(+)-dependent GAPDH and NADH-dependent BDH in B. amyloliquefaciens. Excess GAPDH reduced the fermentation time, increased the 2,3-BD yield by 12.7%, and decreased the acetoin titer by 44.3%. However, the process also enhanced lactate and succinate production. Excess BDH increased the 2,3-BD yield by 16.6% while decreasing acetoin, lactate and succinate production, but prolonged the fermentation time. When BDH and GAPDH were co-overproduced in B. amyloliquefaciens, the fermentation time was reduced. Furthermore, in the NADH-dependent pathways, the molar yield of 2,3-BD was increased by 22.7%, while those of acetoin, lactate and succinate were reduced by 80.8%, 33.3% and 39.5%, relative to the parent strain. In fed-batch fermentations, the 2,3-BD concentration was maximized at 132.9 g/l after 45 h, with a productivity of 2.95 g/l·h.Conclusions/significanceCo-overexpression of bdh and gapA genes proved an effective method for enhancing 2,3-BD production and inhibiting the accumulation of unwanted by-products (acetoin, lactate and succinate). To our knowledge, we have attained the highest 2,3-BD fermentation yield thus far reported for safe microorganisms.

Project description:Background2,3-Butanediol (BD) is a largely used fossil-based platform chemical. The yield and productivity of bio-based BD fermentative production must be increased and cheaper substrates need to be identified, to make bio-based BD production more competitive. As BD bioproduction occurs under microaerobic conditions, a fine tuning and control of the oxygen transfer rate (OTR) is crucial to maximize BD yield and productivity. Very few studies on BD bioproduction focused on the use of non-pathogenic microorganisms and of byproducts as substrate. The goal of this work was to optimize BD bioproduction by the non-pathogenic strain Bacillus licheniformis ATCC9789 by (i) identifying the ranges of volumetric and biomass-specific OTR that maximize BD yield and productivity using standard sugar and protein sources, and (ii) performing a preliminary evaluation of the variation in process performances and cost resulting from the replacement of glucose with molasses, and beef extract/peptone with chicken meat and bone meal, a byproduct of the meat production industry.ResultsOTR optimization with an expensive, standard medium containing glucose, beef extract and peptone revealed that OTRs in the 7-15 mmol/L/h range lead to an optimal BD yield (0.43 ± 0.03 g/g) and productivity (0.91 ± 0.05 g/L/h). The corresponding optimal range of biomass-specific OTR was equal to 1.4-7.9 [Formula: see text], whereas the respiratory quotient ranged from 1.8 to 2.5. The switch to an agro-industrial byproduct-based medium containing chicken meat and bone meal and molasses led to a 50% decrease in both BD yield and productivity. A preliminary economic analysis indicated that the use of the byproduct-based medium can reduce by about 45% the BD production cost.ConclusionsA procedure for OTR optimization was developed and implemented, leading to the identification of a range of biomass-specific OTR and respiratory quotient to be used for the scale-up and control of BD bioproduction by Bacillus licheniformis. The switch to a byproduct-based medium led to a relevant decrease in BD production cost. Further research is needed to optimize the process of BD bioproduction from the tested byproduct-based medium.

Project description:Volatile compounds, such as short chain alcohols, acetoin, and 2,3-butanediol, produced by certain strains of root-associated bacteria (rhizobacteria) elicit induced systemic resistance in plants. The effects of bacterial volatile compounds (BVCs) on plant and fungal growth have been extensively studied; however, the impact of bacterial BVCs on bacterial growth remains poorly understood. In this study the effects of a well-characterized bacterial volatile, 2,3-butanediol, produced by the rhizobacterium Bacillus subtilis, were examined in the rhizosphere. The nature of 2,3-butanediol on bacterial cells was assessed, and the effect of the molecule on root colonization was also determined. Pepper roots were inoculated with three B. subtilis strains: the wild type, a 2,3-butanediol overexpressor, and a 2,3-butanediol null mutant. The B. subtilis null strain was the first to be eliminated in the rhizosphere, followed by the wild-type strain. The overexpressor mutant was maintained at roots for the duration of the experiment. Rhizosphere colonization by a saprophytic fungus declined from 14 days post-inoculation in roots treated with the B. subtilis overexpressor strain. Next, exudates from roots exposed to 2,3-butanediol were assessed for their impact on fungal and bacterial growth in vitro. Exudates from plant roots pre-treated with the 2,3-butanediol overexpressor were used to challenge various microorganisms. Growth was inhibited in a saprophytic fungus (Trichoderma sp.), the 2,3-butanediol null B. subtilis strain, and a soil-borne pathogen, Ralstonia solanacearum. Direct application of 2,3-butanediol to pepper roots, followed by exposure to R. solanacearum, induced expression of Pathogenesis-Related (PR) genes such as CaPR2, CaSAR8.2, and CaPAL. These results indicate that 2,3-butanediol triggers the secretion of root exudates that modulate soil fungi and rhizosphere bacteria. These data broaden our knowledge regarding bacterial volatiles in the rhizosphere and their roles in bacterial fitness and as important inducers of plant defenses.

Dataset Information

A newly isolated Bacillus licheniformis strain thermophilically produces 2,3-butanediol, a platform and fuel bio-chemical.

Publications

A newly isolated Bacillus licheniformis strain thermophilically produces 2,3-butanediol, a platform and fuel bio-chemical.

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