Structural Analysis of Gluco-Oligosaccharides Produced by Leuconostoc lactis and Their Prebiotic Effect

Lee, Sulhee; Park, Jisun; Jang, Jae-Kweon; Yoo, Sang‐Ho

doi:10.3390/molecules24213998

Cited by 15 publications

(16 citation statements)

References 28 publications

(31 reference statements)

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“…Besides, the consumption of these sugars has also been related to a reduction in the risk of suffering colon cancer and cardiovascular diseases, or with the improvement of mineral absorption and immune system response. [ 12 , 13 ]. Due to their techno-functional properties, bioactive oligosaccharides have applications in food technology as bulking and moisture retaining agents, fat and sugar substitutes, textural enhancers, and non-cariogenic ingredients, which give them great biotechnological utility [ 14 , 15 ].…”

Section: Introductionmentioning

confidence: 99%

Molecular characterization and heterologous expression of two α-glucosidases from Metschnikowia spp, both producers of honey sugars

et al. 2020

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Background: α-Glucosidases are widely distributed enzymes with a varied substrate specificity that are traditionally used in biotechnological industries based on oligo-and polysaccharides as starting materials. According to amino acid sequence homology, α-glucosidases are included into two major families, GH13 and GH31. The members of family GH13 contain several α-glucosidases with confirmed hydrolytic activity on sucrose. Previously, a sucrose splitting activity from the nectar colonizing yeast Metschnikowia reukaufii which produced rare sugars with α-(1→1), α-(1→3) and α-(1→6) glycosidic linkages from sucrose was described. Results: In this study, genes codifying for α-glucosidases from the nectaries yeast M. gruessii and M. reukaufii were characterised and heterologously expressed in Escherichia coli for the first time. Recombinant proteins (Mg-αGlu and Mr-αGlu) were purified and biochemically analysed. Both enzymes mainly displayed hydrolytic activity towards sucrose, maltose and p-nitrophenyl-α-d-glucopyranoside. Structural analysis of these proteins allowed the identification of common features from the α-amylase family, in particular from glycoside hydrolases that belong to family GH13. The three acidic residues comprising the catalytic triad were identified and their relevance for the protein hydrolytic mechanism confirmed by site-directed mutagenesis. Recombinant enzymes produced oligosaccharides naturally present in honey employing sucrose as initial substrate and gave rise to mixtures with the same products profile (isomelezitose, trehalulose, erlose, melezitose, theanderose and esculose) previously obtained with M. reukaufii cell extracts. Furthermore, the same enzymatic activity was detected with its orthologous Mg-αGlu from M. gruessii. Interestingly, the isomelezitose amounts obtained in reactions mediated by the recombinant proteins, ~ 170 g/L, were the highest reported so far. Conclusions: Mg/Mr-αGlu were heterologously overproduced and their biochemical and structural characteristics analysed. The recombinant α-glucosidases displayed excellent properties in terms of mild reaction conditions, in addition to pH and thermal stability. Besides, the enzymes produced a rare mixture of hetero-gluco-oligosaccharides by transglucosylation, mainly isomelezitose and trehalulose. These compounds are natural constituents of honey which purification from this natural source is quite unviable, what make these enzymes very interesting for the

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Section: Introductionmentioning

confidence: 99%

Molecular characterization and heterologous expression of two α-glucosidases from Metschnikowia spp, both producers of honey sugars

et al. 2020

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“…Oligosaccharides produced by Leu. lactis SBC001 were purified as described in a previous study [ 17 ]. The culture supernatant was concentrated 10-fold at 60 °C using a rotary evaporator (SB-1200, EYELA, Miyagi, Japan), and the concentrate was loaded onto Bio-gel P2 resin (Bio-Rad Laboratories, Inc., Hercules, CA, USA) packed in a glass Econo-Column (1.5 × 120 cm, Bio-Rad Laboratories, Inc.).…”

Section: Methodsmentioning

confidence: 99%

“…Typically, oligosaccharide-producing LAB include the genera Leuconostoc , Weissella , Streptococcus , Lactobacillus , and Oenococcuss ; these LABs synthesize oligosaccharides by the action of glucansucrase (EC 2.4.1.5), a type of glycosyltransferase, using sucrose as substrate. In addition to glucansucrase, mutansucrase (EC 2.4.1.5), alternansucrase (EC 2.4.1.140), and levansucrase (EC 2.4.1.10) synthesize oligosaccharides with different structures [ 15 , 16 , 17 ].…”

Section: Introductionmentioning

confidence: 99%

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Production Optimization, Structural Analysis, and Prebiotic- and Anti-Inflammatory Effects of Gluco-Oligosaccharides Produced by Leuconostoc lactis SBC001

Kim

Jang

2021

Microorganisms

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Leuconostoc lactis SBC001, isolated from chive, produces glucansucrase and synthesizes oligosaccharides through its enzymatic activity. This study was conducted to optimize oligosaccharide production using response surface methodology, analyze the structure of purified oligosaccharides, and investigate the prebiotic effect on 24 bacterial and yeast strains and the anti-inflammatory activity using RAW 264.7 macrophage cells. The optimal conditions for oligosaccharide production were a culture temperature of 30 °C and sucrose and maltose concentrations of 9.6% and 7.4%, respectively. Based on 1H-NMR spectroscopic study, the oligosaccharides were identified as gluco-oligosaccharides that consisted of 23.63% α-1,4 glycosidic linkages and 76.37% α-1,6 glycosidic linkages with an average molecular weight of 1137 Da. The oligosaccharides promoted the growth of bacterial and yeast strains, including Lactobacillus plantarum, L. paracasei, L. johnsonii, Leuconostoc mesenteroides, L. rhamnosus, and Saccharomyces cerevisiae. When lipopolysaccharide-stimulated RAW 264.7 cells were treated with the oligosaccharides, the production of nitric oxide was decreased; the expression of inducible nitric oxide synthase, tumor necrosis factor-α, interleukin (IL)-1β, IL-6, and IL-10 was suppressed; and the nuclear factor-kappa B signaling pathway was inhibited. In conclusion, the gluco-oligosaccharides obtained from Leu. lactis SBC001 exhibited a prebiotic effect on six bacterial and yeast strains and anti-inflammatory activity in RAW 264.7 macrophage cells.

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“…Relaxation times, including longitudinal spin-lattice relaxation (T 1 ) and transverse spin-spin relaxation (T 2 ), reveal valuable information for studying molecular dynamics in practical NMR applications [1][2][3]. The combination of relaxation times and other detection parameters, such as chemical shifts and spin-spin scalar (J) couplings, endows NMR spectroscopy with its superiority over other detection methods in structural studies [4][5][6]. In detail, T 1 relaxation describes the recovery process of magnetizations along the longitudinal direction after signal excitation.…”

Section: Introductionmentioning

confidence: 99%

NMR Relaxation Measurements on Complex Samples Based on Real-Time Pure Shift Techniques

Lin

Zhan

et al. 2020

Molecules

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Longitudinal spin-lattice relaxation (T1) and transverse spin-spin relaxation (T2) reveal valuable information for studying molecular dynamics in NMR applications. Accurate relaxation measurements from conventional 1D proton spectra are generally subject to challenges of spectral congestion caused by J coupling splittings and spectral line broadenings due to magnetic field inhomogeneity. Here, we present an NMR relaxation method based on real-time pure shift techniques to overcome these two challenges and achieve accurate measurements of T1 and T2 relaxation times from complex samples that contain crowded NMR resonances even under inhomogeneous magnetic fields. Both theoretical analyses and detailed experiments are performed to demonstrate the effectiveness and ability of the proposed method for accurate relaxation measurements on complex samples and its practicability to non-ideal magnetic field conditions.

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Structural Analysis of Gluco-Oligosaccharides Produced by Leuconostoc lactis and Their Prebiotic Effect

Cited by 15 publications

References 28 publications

Molecular characterization and heterologous expression of two α-glucosidases from Metschnikowia spp, both producers of honey sugars

Molecular characterization and heterologous expression of two α-glucosidases from Metschnikowia spp, both producers of honey sugars

Production Optimization, Structural Analysis, and Prebiotic- and Anti-Inflammatory Effects of Gluco-Oligosaccharides Produced by Leuconostoc lactis SBC001

NMR Relaxation Measurements on Complex Samples Based on Real-Time Pure Shift Techniques

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