Novel β-galactosynthase–β-mannosynthase dual activity of β-galactosidase from Aspergillus oryzae uncovered using monomer sugar substrates

Tan, Roger Salvacion; Hinou, Hiroshi; Nishimura, S.‐I.

doi:10.1039/c6ra08060j

Cited by 6 publications

(4 citation statements)

References 20 publications

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“…In addition to glycosidic bond cleavage, glycoside hydrolases (GHs) can be used for the synthesis of glycoside bonds in vitro via reverse hydrolysis reaction without the need for cofactors, such as uridine diphosphate (Lu et al 2015;Perugino et al 2004;Rosengren et al 2019;Xu et al 2016). Recently, several glycosidases, such as β-glucosidase, α-glucosidase, α-L -rhamnosidase, β-mannosidase and β-galactosidase, have been used to synthesize glycosides via the reverse hydrolysis reaction (Lu et al 2015;Arthornthurasuk et al 2018;Chen et al 2019;Ajisaka et al 2012;Tan et al 2016). Gentiooligosaccharides, octyl glucoside, laminaribiose, sophorose, cellobiose, and gentiobiose have been synthesized by β-glucosidase via reverse hydrolysis reaction (Semenova et al 2015;Arthornthurasuk et al 2018;Wang et al 2018;Ravet et al 1993).…”

Section: Introductionmentioning

confidence: 99%

A novel strategy for efficient disaccharides synthesis from glucose by β-glucosidase

Niu

Liu

Feng

et al. 2020

Bioresour. Bioprocess.

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Oligosaccharides have important therapeutic applications. A useful route for oligosaccharides synthesis is reverse hydrolysis by β-glucosidase. However, the low conversion efficiency of disaccharides from monosaccharides limits its large-scale production because the equilibrium is biased in the direction of hydrolysis. Based on the analysis of the docking results, we hypothesized that the hydropathy index of key amino acid residues in the catalytic site is closely related with disaccharide synthesis and more hydrophilic residues located in the catalytic site would enhance reverse hydrolysis activity. In this study, positive variants TrCel1b I177S , TrCel1b I177S/I174S , and TrCel1b I177S/I174S/W173H , and one negative variant TrCel1b N240I were designed according to the Hydropathy Index For Enzyme Activity (HIFEA) strategy. The reverse hydrolysis with TrCel1b I177S/I174S/W173H was accelerated and then the maximum total production (195.8 mg/mL/ mg enzyme) of the synthesized disaccharides was increased by 3.5-fold compared to that of wild type. On the contrary, TrCel1b N240I lost reverse hydrolysis activity. The results demonstrate that the average hydropathy index of the key amino acid residues in the catalytic site of TrCel1b is an important factor for the synthesis of laminaribiose, sophorose, and cellobiose. The HIFEA strategy provides a new perspective for the rational design of β-glucosidases used for the synthesis of oligosaccharides.

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Section: Introductionmentioning

confidence: 99%

A novel strategy for efficient disaccharides synthesis from glucose by β-glucosidase

Niu

Liu

Feng

et al. 2020

Bioresour. Bioprocess.

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“…The acceptability of the free galactose as substrate of Ao-β-gal has been reported recently (Tan et al, 2016). Ao-β-gal has been observed to synthesized galactobiose in Gal-β(1-6)-Gal glycosydic linkage after 24 hours of incubation.…”

Section: Gos Production and Quantitationmentioning

confidence: 88%

“…Another novel activity of Ao-β-gal is its reported β-mannosynthase (Tan et al, 2016). In a reported reaction of the enzyme with the monosaccharide mannose, as the substrate, Ao-β-gal was able to synthesize mannobiose in Man-β(1-6)-Man glycosydic linkage after 24 hours of incubation.…”

Section: Mos Production and Quantitationmentioning

confidence: 99%

“…Recently, we reported a novel β-galactosynthase -β-mannosynthase dual-activity of Ao-β-gal, which was uncovered for the first time using free monosaccharide substrates (Tan et al, 2016). The enzyme was found to have the capacity to synthesize galactobiose and mannobiose from monosaccharides galactose and mannose, respectively, which were both linked in a β(1-6) fashion.…”

Section: Introductionmentioning

confidence: 99%

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Modulation of cancer cell proliferation by unusually produced β(1-6) linked Mannan-oligosaccharides and β(1-6) linked Galactooligosaccharides using β–galactosidase from Aspergillus oryzae

Tan

2018

Int. j. adv. appl. sci.

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This study introduced an alternative way to produce NDOs, utilizing the recently reported unusual β-galactosynthase-β-mannosynthase dual activity of this enzyme, using free monosaccharide substrates. The enzyme successfully converted galactose and mannose monomer sugars efficiently to NDOs. Glycoblotting and MALDI-TOFMS analyses confirmed that the enzyme produced a maximum of 46.37% galacto-oligosaccharides (36.88% galactobiose, 8.52 % galactotriose, and 0.9746 % galactotetraose) from galactose monosaccharides and 32.62 % mannan-oligosaccharides (30.95 % mannobiose and 1.68 % mannotriose) from mannose monosaccharides. Both MOS and GOS modulated cell growth of different human cancer cell lines, with apparent antiproliferative effect on prostate adenocarcinoma (PC-3) cells and mitogenic effects on A549, HepG2, and HT-29. Mitochondrial membrane potential and careful morphological evaluation of PC-3 cells treated with GOS and MOS suggested that MOS treated cells showed possible inhibition of cellular adhesion that might have caused apoptosis to advance. The high-yielding NDOs synthase character of this cheaply available βgalactosidase makes this new strategy a good potential alternative in producing prebiotic NDOs. Keywords:Non-digestible oligosaccharides Galacto-oligosaccharides Mannan-oligosaccharides Asprgillus oryzae β-galactosidase Glycoblotting

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Analysis of Carbohydrates and Glycoconjugates by Matrix‐assisted Laser Desorption/Ionization Mass Spectrometry: An Update for 2015–2016

Harvey

2021

Mass Spectrometry Reviews

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This review is the ninth update of the original article published in 1999 on the application of matrix‐assisted laser desorption/ionization (MALDI) mass spectrometry to the analysis of carbohydrates and glycoconjugates and brings coverage of the literature to the end of 2016. Also included are papers that describe methods appropriate to analysis by MALDI, such as sample preparation techniques, even though the ionization method is not MALDI. Topics covered in the first part of the review include general aspects such as theory of the MALDI process, matrices, derivatization, MALDI imaging, fragmentation and arrays. The second part of the review is devoted to applications to various structural types such as oligo‐ and poly‐saccharides, glycoproteins, glycolipids, glycosides and biopharmaceuticals. Much of this material is presented in tabular form. The third part of the review covers medical and industrial applications of the technique, studies of enzyme reactions and applications to chemical synthesis. The reported work shows increasing use of combined new techniques such as ion mobility and the enormous impact that MALDI imaging is having. MALDI, although invented over 30 years ago is still an ideal technique for carbohydrate analysis and advancements in the technique and range of applications show no sign of deminishing. © 2020 Wiley Periodicals, Inc.

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Novel β-galactosynthase–β-mannosynthase dual activity of β-galactosidase from Aspergillus oryzae uncovered using monomer sugar substrates

Abstract: We uncovered β-galactosynthase–β-mannosynthase dual-activity of β-galactosidase (A. oryzae) that could revolutionize chemoenzymatic glycan and NDOs syntheses.

Cited by 6 publications

References 20 publications

A novel strategy for efficient disaccharides synthesis from glucose by β-glucosidase

A novel strategy for efficient disaccharides synthesis from glucose by β-glucosidase

Modulation of cancer cell proliferation by unusually produced β(1-6) linked Mannan-oligosaccharides and β(1-6) linked Galactooligosaccharides using β–galactosidase from Aspergillus oryzae

Analysis of Carbohydrates and Glycoconjugates by Matrix‐assisted Laser Desorption/Ionization Mass Spectrometry: An Update for 2015–2016

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