Isolation and characterization of novel 1,3-propanediol-producing Lactobacillus panis PM1 from bioethanol thin stillage

Khan, Nurul H.; Kang, Tae Seok; Grahame, Douglas A. S.; Haakensen, Monique; Ratanapariyanuch, Kornsulee; Reaney, Martin J. T.; Korber, Darren R.; Tanaka, Takuji

doi:10.1007/s00253-012-4386-4

Cited by 38 publications

(46 citation statements)

References 31 publications

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“…Overall, the genes of the PM1 strain showed higher similarities with those of group III lactobacilli (64 to 88%) than those of group I and II lactobacilli (42 to 73%), and the sequence of the L. reuteri JCM 1112 strain showed the highest similarity (64, 83, and 88% in the TPI, FBA, and PGI genes, respectively) with that of the PM1 strain for all genes except the FK gene (28 and 30% to the two FK genes of the L. reuteri JCM 1112 strain). Our previous phylogenetic work showed higher identity (96%) in 16S rRNA sequences to the 16S rRNA sequence of L. reuteri (7). Combined with the present results, this indicates that L. panis PM1 and L. reuteri are close relatives.…”

Section: Figsupporting

confidence: 73%

“…Lactobacillus panis PM1 has been described as a group III lactobacillus that uses the 6-PG/PK and various external electron acceptor pathways (e.g., glycerol, citrate, and oxygen) for its central metabolism and NADH recycling system (7)(8)(9), and its inability to grow on fructose medium has been reported by our group (7). In this study, we isolated a variant PM1 strain able to grow on fructose as the sole carbon source.…”

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confidence: 85%

“…5). Generally, DHAP is formed by the EM or glycerol oxidative (glycerol-to-DHAP) pathway; however, L. panis PM1 does not possess the glycerol oxidative routes (7,8). Thus, the accumulation of DHAP suggested the utilization of the EM pathway and provided a direct reason for growth cessation with fructose but not with glucose.…”

Section: Figmentioning

confidence: 99%

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Regulation of Dual Glycolytic Pathways for Fructose Metabolism in Heterofermentative Lactobacillus panis PM1

Kang

Korber

Tanaka

2013

Appl Environ Microbiol

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Lactobacillus panis PM1 belongs to the group III heterofermentative lactobacilli that use the 6-phosphogluconate/phosphoketolase (6-PG/PK) pathway as their central metabolic pathway and are reportedly unable to grow on fructose as a sole carbon source. We isolated a variant PM1 strain capable of sporadic growth on fructose medium and observed its distinctive characteristics of fructose metabolism. The end product pattern was different from what is expected in typical group III lactobacilli using the 6-PG/PK pathway (i.e., more lactate, less acetate, and no mannitol). In addition, in silico analysis revealed the presence of genes encoding most of critical enzymes in the Embden-Meyerhof (EM) pathway. These observations indicated that fructose was metabolized via two pathways. Fructose metabolism in the PM1 strain was influenced by the activities of two enzymes, triosephosphate isomerase (TPI) and glucose 6-phosphate isomerase (PGI). A lack of TPI resulted in the intracellular accumulation of dihydroxyacetone phosphate (DHAP) in PM1, the toxicity of which caused early growth cessation during fructose fermentation. The activity of PGI was enhanced by the presence of glyceraldehyde 3-phosphate (GAP), which allowed additional fructose to enter into the 6-PG/PK pathway to avoid toxicity by DHAP. Exogenous TPI gene expression shifted fructose metabolism from heterolactic to homolactic fermentation, indicating that TPI enabled the PM1 strain to mainly use the EM pathway for fructose fermentation. These findings clearly demonstrate that the balance in the accumulation of GAP and DHAP determines the fate of fructose metabolism and the activity of TPI plays a critical role during fructose fermentation via the EM pathway in L. panis PM1. L actobacilli generate metabolic energy (i.e., ATP) mainly by substrate-level phosphorylation by relatively simple pathways: the Embden-Meyerhof (EM) pathway and/or the 6-phosphogluconate/phosphoketolase (6-PG/PK) pathway. The members of the genus Lactobacillus are subdivided into three groups according to their fermentative characteristics on the basis of the presence of these pathways (1, 2). Group I lactobacilli are obligatory homofermentative lactobacilli that exclusively ferment hexose sugars (e.g., glucose) to lactate via the EM pathway; however, they do not metabolize pentose sugars or gluconate. Group II lactobacilli are the facultative heterofermentative lactobacilli that ferment hexoses to lactate via the EM pathway. They can also ferment pentose sugars using an inducible phosphoketolase, producing lactate and acetate, and they can yield carbon dioxide from gluconate but not from glucose. Group III lactobacilli are the obligatory heterofermentative lactobacilli that ferment hexoses to lactate, ethanol and/or acetate, and carbon dioxide, which is a distinct feature of the group III lactobacilli. Group III lactobacilli exclusively use the 6-PG/PK pathway to achieve this metabolism.Fructose, an abundant hexose sugar found in many plants, is one of the main monosaccharides for bacteria...

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Section: Figsupporting

confidence: 73%

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confidence: 85%

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Regulation of Dual Glycolytic Pathways for Fructose Metabolism in Heterofermentative Lactobacillus panis PM1

Kang

Korber

Tanaka

2013

Appl Environ Microbiol

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“…Interestingly, strain KF147 was previously reported as being unable to metabolize glycerol (37). Other LAB were shown to degrade glycerol (51,52), and therefore the capacity to consume this compound might depend on the environmental context and other nutrients to which the cells are exposed.…”

Section: Discussionmentioning

confidence: 99%

Lactococcus lactis Metabolism and Gene Expression during Growth on Plant Tissues

Golomb

Marco

2014

J. Bacteriol.

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Lactic acid bacteria have been isolated from living, harvested, and fermented plant materials; however, the adaptations these bacteria possess for growth on plant tissues are largely unknown. In this study, we investigated plant habitat-specific traits of Lactococcus lactis during growth in an Arabidopsis thaliana leaf tissue lysate (ATL). L. lactis KF147, a strain originally isolated from plants, exhibited a higher growth rate and reached 7.9-fold-greater cell densities during growth in ATL than the dairy-associated strain L. lactis IL1403. Transcriptome profiling (RNA-seq) of KF147 identified 853 induced and 264 repressed genes during growth in ATL compared to that in GM17 laboratory culture medium. Genes induced in ATL included those involved in the arginine deiminase pathway and a total of 140 carbohydrate transport and metabolism genes, many of which are involved in xylose, arabinose, cellobiose, and hemicellulose metabolism. The induction of those genes corresponded with L. lactis KF147 nutrient consumption and production of metabolic end products in ATL as measured by gas chromatography-time of flight mass spectrometry (GC-TOF/MS) untargeted metabolomic profiling. To assess the importance of specific plant-inducible genes for L. lactis growth in ATL, xylose metabolism was targeted for gene knockout mutagenesis. Wild-type L. lactis strain KF147 but not an xylA deletion mutant was able to grow using xylose as the sole carbon source. However, both strains grew to similarly high levels in ATL, indicating redundancy in L. lactis carbohydrate metabolism on plant tissues. These findings show that certain strains of L. lactis are well adapted for growth on plants and possess specific traits relevant for plant-based food, fuel, and feed fermentations. L actic acid bacteria (LAB) found on plants are essential for the production of a wide variety of plant-derived fermented foods with desirable organoleptic properties, improved nutritional attributes, and extended shelf life (1). LAB are a diverse group of bacterial species in the Firmicutes phylum that are characterized by the production of lactic acid as the main end product of carbohydrate metabolism. Some of the most commonly recognized genera among the LAB include Lactococcus, Lactobacillus, Leuconostoc, Pediococcus, and Oenococcus. LAB-fermented plantbased food products include sauerkraut, sourdough, and olives, as well as diverse foods unique to different ethnic groups (e.g., pulque, nukadoko, or gundruk [2,3]). LAB are responsible for the conversion of plant tissues such as alfalfa into silage for animal feed with enhanced nutritional content and stability (4). LAB are also used to ferment plant-based substrates into industrial-grade lactic acid for the manufacture of polylactide bioplastics (5). Conversely, LAB are frequent contaminants of bioethanol fermentations (6).In contrast to commercial dairy fermentations that utilize starter cultures, initiation of plant-based fermentations typically relies on the LAB that are located on or associated with plant tis...

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“…Because of their small genomes and relatively simple metabolic pathways, lactobacilli have been intensively utilized as cell factories for the production of platform chemicals, including lactic acid, 2,3-butanediol, 1,3-PDO, ethanol, food flavoring agents, sweeteners, and complex polysaccharides, through metabolic engineering approaches (8). Lactobacillus panis PM1 is a natural 1,3-PDO-producing organism originally isolated from TS (9). This strain achieves primary carbohydrate fermentation through substrate-level phosphorylation via the 6-phosphogluconate/phosphoketolase (6-PG/ PK) pathway wherein NADH is mainly recycled back to NAD ϩ through the production of lactic acid and ethanol (Fig.…”

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confidence: 99%

Metabolic Engineering of a Glycerol-Oxidative Pathway in Lactobacillus panis PM1 for Utilization of Bioethanol Thin Stillage: Potential To Produce Platform Chemicals from Glycerol

Kang

Korber

Tanaka

2014

Appl Environ Microbiol

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Lactobacillus panis PM1 has the ability to produce 1,3-propanediol (1,3-PDO) from thin stillage (TS), which is the major waste material after bioethanol production, and is therefore of significance. However, the fact that L. panis PM1 cannot use glycerol as a sole carbon source presents a considerable problem in terms of utilization of this strain in a wide range of industrial applications. Accordingly, L. panis PM1 was genetically engineered to directly utilize TS as a fermentable substrate for the production of valuable platform chemicals without the need for exogenous nutrient supplementation (e.g., sugars and nitrogen sources). An artificial glycerol-oxidative pathway, comprised of glycerol facilitator, glycerol kinase, glycerol 3-phosphate dehydrogenase, triosephosphate isomerase, and NADPH-dependent aldehyde reductase genes of Escherichia coli, was introduced into L. panis PM1 in order to directly utilize glycerol for the production of energy for growth and value-added chemicals. A pH 6.5 culture converted glycerol to mainly lactic acid (85.43 mM), whereas a significant amount of 1,3-propanediol (59.96 mM) was formed at pH 7.5. Regardless of the pH, ethanol (82.16 to 83.22 mM) was produced from TS fermentations, confirming that the artificial pathway metabolized glycerol for energy production and converted it into lactic acid or 1,3-PDO and ethanol in a pH-dependent manner. This study demonstrates the cost-effective conversion of TS to value-added chemicals by the engineered PM1 strain cultured under industrial conditions. Thus, application of this strain or these research findings can contribute to reduced costs of bioethanol production.T hin stillage (TS) is a major fermentation residue from the dry-grind process of bioethanol production. Its downstream treatment, including making a condensed form for animal feed, is an energy-and cost-intensive process (1), while its utilization as animal feed is hardly a high-value application of this industrial by-product and hence does not significantly reduce the cost of ethanol production. TS contains a variety of complex nutrients, including various carbohydrates, minerals, and amino acids. Glycerol (up to 2%) is generated in TS during bioethanol production regardless of feedstock (i.e., sugar cane or corn) (2). Glycerol is at a higher reduced state than carbohydrates (i.e., fermentable sugars). The reduced nature of glycerol allows synthesis of fuels and other reduced chemicals at higher yields with minimal by-products compared to common sugars (e.g., glucose and xylose); however, the high degree of reduction in glycerol makes it difficult for microorganisms to utilize under anaerobic conditions. A few bacteria, including Klebsiella, Enterobacter, and Clostridium, have been exploited to bioconvert glycerol to industrially relevant chemicals (e.g., 1,3-propanediol [1,3-PDO], 3-hydroxypropionic acid, butanol, and ethanol) (3-7). Since these organisms are classified as opportunistic pathogens, industrial applications of these strains may pose issues, particularly re...

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Isolation and characterization of novel 1,3-propanediol-producing Lactobacillus panis PM1 from bioethanol thin stillage

Cited by 38 publications

References 31 publications

Regulation of Dual Glycolytic Pathways for Fructose Metabolism in Heterofermentative Lactobacillus panis PM1

Regulation of Dual Glycolytic Pathways for Fructose Metabolism in Heterofermentative Lactobacillus panis PM1

Lactococcus lactis Metabolism and Gene Expression during Growth on Plant Tissues

Metabolic Engineering of a Glycerol-Oxidative Pathway in Lactobacillus panis PM1 for Utilization of Bioethanol Thin Stillage: Potential To Produce Platform Chemicals from Glycerol

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