Purification and characterization of a novel glucansucrase from Leuconostoc lactis EG001

Kim, Yongmo; Yeon, Min Ji; Choi, Nack-Shick; Chang, Young-Hyo; Jung, Min Young; Song, Jae Jun; Kim, Joong Su

doi:10.1016/j.micres.2009.08.005

Cited by 23 publications

(21 citation statements)

References 20 publications

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“…Complete genome sequence of L. mesenteroides ATCC 8293 (GenBank no. NC_008531), a strain equivalent to KCTC 3718 [29], was used as a reference sequence to detect genetic alterations. Conventional PCR and Sanger sequencing service (Macrogen, Korea) were used to confirm the sequence changes observed after Illumina sequencing.…”

Section: Methodsmentioning

confidence: 99%

Long-term adaptive evolution of Leuconostoc mesenteroides for enhancement of lactic acid tolerance and production

Kim

Lee

2016

Biotechnol Biofuels

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BackgroundLactic acid has been approved by the United States Food and Drug Administration as Generally Regarded As Safe (GRAS) and is commonly used in the cosmetics, pharmaceutical, and food industries. Applications of lactic acid have also emerged in the plastics industry. Lactic acid bacteria (LAB), such as Leuconostoc and Lactobacillus, are widely used as lactic acid producers for food-related and biotechnological applications. Nonetheless, industrial mass production of lactic acid in LAB is a challenge mainly because of growth inhibition caused by the end product, lactic acid. Thus, it is important to improve acid tolerance of LAB to achieve balanced cell growth and a high titer of lactic acid. Recently, adaptive evolution has been employed as one of the strategies to improve the fitness and to induce adaptive changes in bacteria under specific growth conditions, such as acid stress.ResultsWild-type Leuconostoc mesenteroides was challenged long term with exogenously supplied lactic acid, whose concentration was increased stepwise (for enhancement of lactic acid tolerance) during 1 year. In the course of the adaptive evolution at 70 g/L lactic acid, three mutants (LMS50, LMS60, and LMS70) showing high specific growth rates and lactic acid production were isolated and characterized. Mutant LMS70, isolated at 70 g/L lactic acid, increased d-lactic acid production up to 76.8 g/L, which was twice that in the wild type (37.8 g/L). Proteomic, genomic, and physiological analyses revealed that several possible factors affected acid tolerance, among which a mutation of ATPase ε subunit (involved in the regulation of intracellular pH) and upregulation of intracellular ammonia, as a buffering system, were confirmed to contribute to the observed enhancement of tolerance and production of d-lactic acid.ConclusionsDuring adaptive evolution under lethal stress conditions, the fitness of L. mesenteroides gradually increased to accumulate beneficial mutations according to the stress level. The enhancement of acid tolerance in the mutants contributed to increased production of d-lactic acid. The observed genetic and physiological changes may systemically help remove protons and retain viability at high lactic acid concentrations.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0662-3) contains supplementary material, which is available to authorized users.

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Section: Methodsmentioning

confidence: 99%

Long-term adaptive evolution of Leuconostoc mesenteroides for enhancement of lactic acid tolerance and production

Kim

Lee

2016

Biotechnol Biofuels

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“…Glucansucrase enzymes are interesting biocatalysts for the glycosylation of small organic molecules, because they are promiscuous towards a wide range of acceptor substrates (Monsan et al 2010; Leemhuis et al 2012). Using inexpensive sucrose as glycosyl donor, glucansucrase enzymes glucosylate, for instance, phenolic compounds such as catechol (Meulenbeld and Hartmans 2000; Devlamynck et al 2016; te Poele et al 2016) and hydroquinone (Seo et al 2009; Devlamynck et al 2016), primary alcohols (Seibel et al 2006; Devlamynck et al 2016), and l -ascorbic acid (Kim et al 2010). Glycosylation may change the physico-chemical and biological properties of molecules, enhance bioavailability of antibiotics, or improve stability of molecules against auto-oxidation (Desmet et al 2012).…”

Section: Introductionmentioning

confidence: 99%

Catechol glucosides act as donor/acceptor substrates of glucansucrase enzymes of Lactobacillus reuteri

Poele

Valk

Devlamynck

et al. 2017

Appl Microbiol Biotechnol

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Previously, we have shown that the glucansucrase GtfA-ΔN enzyme of Lactobacillus reuteri 121, incubated with sucrose, efficiently glucosylated catechol and we structurally characterized catechol glucosides with up to five glucosyl units attached (te Poele et al. in Bioconjug Chem 27:937–946, 2016). In the present study, we observed that upon prolonged incubation of GtfA-ΔN with 50 mM catechol and 1000 mM sucrose, all catechol had become completely glucosylated and then started to reappear. Following depletion of sucrose, this glucansucrase GtfA-ΔN used both α-d-Glcp-catechol and α-d-Glcp-(1→4)-α-d-Glcp-catechol as donor substrates and transferred a glucose unit to other catechol glycoside molecules or to sugar oligomers. In the absence of sucrose, GtfA-ΔN used α-d-Glcp-catechol both as donor and acceptor substrate to synthesize catechol glucosides with 2 to 10 glucose units attached and formed gluco-oligosaccharides up to a degree of polymerization of 4. Also two other glucansucrases tested, Gtf180-ΔN from L. reuteri 180 and GtfML1-ΔN from L. reuteri ML1, used α-d-Glcp-catechol and di-glucosyl-catechol as donor/acceptor substrate to synthesize both catechol glucosides and gluco-oligosaccharides. With sucrose as donor substrate, the three glucansucrase enzymes also efficiently glucosylated the phenolic compounds pyrogallol, resorcinol, and ethyl gallate; also these mono-glucosides were used as donor/acceptor substrates.Electronic supplementary materialThe online version of this article (doi:10.1007/s00253-017-8190-z) contains supplementary material, which is available to authorized users.

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“…This research demonstrated that up to 0.3% β-glucan could be added to yogurt, which meets the nutrient guidelines for added nutritional benefits, without affecting the characteristics of yogurt significantly. The solidification of milk using glucansucrase produced by Leuconostoc species was previously observed by Kim et al [18,19] when the W. hellenica SKKimchi3 strain and Lac. lactis EG001 were cultivated in an MRS medium where 10% sucrose was added to skim milk.…”

Section: Application Of the Dsrlm34 For The Solidification Of Sucrosementioning

confidence: 63%

“…lactis EG001, showing a difference of 30 amino acids from the EG001 glucansucrase amino acid sequence (Fig. 2) [19].…”

Section: Cloning and Nucleotide Sequence Determination Of Glucansucrasementioning

confidence: 99%

“…However, the solidification of semi-skimmed milk using glucansucrases has an advantage in minimizing the heat-induced changes and the starter and lactic acid bacteria survival during drying. Few studies on exopolysaccharide produced by lactic acid bacteria isolated from kimchi have been reported [18][19][20]. However, food industry studies on the solidification of milk using glucansucrase produced by Leuconostoc species isolated from kimchi have rarely been reported.…”

Section: Introductionmentioning

confidence: 99%

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Molecular cloning and characterization of a novel glucansucrase from Leuconostoc mesenteroides subsp. mesenteroides LM34

Kang

Nguyen

Jeong

et al. 2014

Biotechnol Bioproc E

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Leuconostoc mesenteroides LM34 was isolated from kimchi, a traditional fermented Korean food. L. mesenteroides LM34 produced extracellular glucansucrase (DSRLM34), which is responsible for the synthesis of soluble glucan using sucrose. The DSRLM34 gene consists of a 4,503 bp open reading frame (ORF) and encodes an enzyme of 1,500 amino acids with an apparent molecular mass of 165 kDa. The deduced amino-acid sequence showed the highest amino-acid sequence identity (98%) to that of glucansucrase of Lactobacillus lactis. The gene was over-expressed in Escherichia coli strain and the recombinant enzyme (rDSRLM34) was purified. Both DSRLM34 and rDSRLM34 synthesized glucan mainly containing α-1, 6 glucosidic linkage and branched α-1, 3 glucosidic linkages. The enzyme exhibited optimum activity at 30 o C and pH 5.0. DSRLM34 has promising potential as a thickening agent in sucrose-supplemented milk.

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Purification and characterization of a novel glucansucrase from Leuconostoc lactis EG001

Cited by 23 publications

References 20 publications

Long-term adaptive evolution of Leuconostoc mesenteroides for enhancement of lactic acid tolerance and production

Long-term adaptive evolution of Leuconostoc mesenteroides for enhancement of lactic acid tolerance and production

Catechol glucosides act as donor/acceptor substrates of glucansucrase enzymes of Lactobacillus reuteri

Molecular cloning and characterization of a novel glucansucrase from Leuconostoc mesenteroides subsp. mesenteroides LM34

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