Biotechnology Letters

, Volume 39, Issue 8, pp 1229–1235 | Cite as

Molecular cloning and expression of a glycosyltransferase from Bacillus subtilis for modification of morin and related polyphenols

  • Qianqian Wang
  • Yixiang Xu
  • Jiaqi Xu
  • Xudong Wang
  • Chen Shen
  • Yan Zhang
  • Xiufeng Liu
  • Boyang Yu
  • Jian ZhangEmail author
Original Research Paper



To characterize glycosyltransferases from Bacillus subtilis ATCC 6633 and investigate their substrate specificity towards plant polyphenols.


Among the cloned and expressed six UDP-glycosyltransferases (BsGT1-6), BsGT-1 showed activity with a wide range of polyphenols: morin, quercetin, alizarin, rehin, curcumin and aloe emodin. The gene of BsGT-1 has an ORF of 1206 bp encoding 402 amino acids. The recombinant enzyme was purified to homogeneity by Ni–NTA affinity chromatograph, and its biochemical characteristics were identified by HPLC–UV/MS, 1H-NMR and 13C-NMR. BsGT-1 has an MW of approx. 46 kDa as indicated by SDS-PAGE; its activity was optimal at 40 °C and pH 8.5. The Km value of BsGT-1 towards morin was 110 μM.


BsGT-1 from B. subtilis was cloned. It had high catalytic capabilities towards polyphenols which would make it feasible for the structural modification of polyphenols.


Bacillus subtilis Glycosyltransferase Glycosylation Morin Polyphenols 


Supporting information

Section 1—The structure elucidation of compounds 1a and 2a.

Section 2—The structure elucidation of compounds 3a, 4a and 6a.

Section 3—The structure elucidation of compound 5a.

Supplementary Table 1—The genebank accession numbers and the primers of BsGT1-6.

Supplementary Fig. 1—HPLC chromatogram of BsGT-1 reaction products of flavonols: (A) compound 1 with BsGT-1. (B) compound 2 with BsGT-1.

Supplementary Fig. 2—HPLC chromatogram of BsGT-1 reaction product of anthraquinone: (A) compound 3 with BsGT-1. (B) compound 4 with BsGT-1. (C) compound 6 with BsGT-1.

Supplementary Fig. 3—HPLC Chromatogram of reaction product of compound 5 with BsGT-1.

Supplementary Fig. 4—Enzymatic activity of BsGT-1 depending in pH (A) and temperature (B).

Supplementary material

10529_2017_2352_MOESM1_ESM.doc (1.2 mb)
Supplementary material 1 (DOC 1232 kb)


  1. Agarwal SK, Singh SS, Verma S, Kumar S (1999) Two new anthraquinone derivatives from Rheum ernodi. Indian J Chem B 38:749–751Google Scholar
  2. Arima H, Danno G (2002) Isolation of antimicrobial compounds from guava (Psidium guajava L.) and their structural elucidation. Biosci Biotechnol Biochem 66:1727–1730CrossRefPubMedGoogle Scholar
  3. Bourne Y, Henrissat B (2001) Glycoside hydrolases and glycosyltransferases: families and functional modules. Curr Opin Struct Biol 11:593–600CrossRefPubMedGoogle Scholar
  4. Bowles D, Isayenkove J, Lim EK, Poppenberger B (2005) Glycosyltransferases: managers of small molecules. Curr Opin Plant Biol 8:254–263CrossRefPubMedGoogle Scholar
  5. Brabcova J, Carrasco-Lopez C, Bavaro T, Hermoso JA, Palomo JM (2014) Escherichia coli LacZ β-galactosidase inhibition by monohydroxyacetylated glycopyranosides: role of the acetyl groups. J Mol Catal B 107:31–38CrossRefGoogle Scholar
  6. Choi SH, Ryu M, Yoon YJ, Kim DM, Lee EY (2012) Glycosylation of various flavonoids by recombinant oleandomycin glycosyltransferase from Streptomyces antibioticus in batch and repeated batch modes. Biotechnol Lett 34:499–505CrossRefPubMedGoogle Scholar
  7. De Bruyn F, De Paepe B, Maertens J, Beauprez J, De Cocker P, Mincke S et al (2015) Development of an in vivo glucosylation platform by coupling production to growth: production of phenolic glucosides by a glycosyltransferase of Vitis vinifera. Biotechnol Bioeng 112:1594–1603CrossRefPubMedGoogle Scholar
  8. Engstrom KM, Daanen JF, Wagaw S, Stewart AO (2006) Gram scale synthesis of the glucuronide metabolite of ABT-724. J Org Chem 71:78–83Google Scholar
  9. Figueroa CM, Asención Diez MD, Kuhn ML, McEwen S, Salerno G, Iglesias AA et al (2013) The unique nucleotide specificity of the sucrose synthase from Thermosynechococcus elongatus. FEBS Lett 587:165–169CrossRefPubMedGoogle Scholar
  10. Frąckowiak A, Skibiński P, Gaweł W, Zaczyńska E, Czarny A, Gancarz R (2010) Synthesis of glycoside derivatives of hydroxyanthraquinone with ability to dissolve and inhibit formation of crystals of calcium oxalate. Potential compounds in kidney stone therapy. Eur J Med Chem 45:1001–1007CrossRefPubMedGoogle Scholar
  11. Gan RY, Deng ZQ, Yan AX, Shah NP, Lui WY, Chan CL, Corke H (2016) Pigmented edible bean coats as natural sources of polyphenols with antioxidant and antibacterial effects. LWT-Food Sci Technol 73:168–177CrossRefGoogle Scholar
  12. Gantt RW, Goff RD, Williams GJ, Thorson JS (2008) Probing the aglycon promiscuity of an engineered glycosyltransferase. Angew Chem Int Ed 47:8889–8892CrossRefGoogle Scholar
  13. Iwakiria T, Masea S, Murakamia T, Matsumotob M, Hamadab H, Nakayamac T, Ozaki S (2013) Glucosylation of hydroxyflavones by glucosyltransferases from Phytolacca americana. J Mol Catal B 90:61–65CrossRefGoogle Scholar
  14. Jiang JR, Yuan S, Ding JF, Zhu SC, Xu HD, Chen T, Cong XD, Xu WP, Ye H, Dai YJ (2008) Conversion of puerarin into its 7-O-glycoside derivatives by Microbacterium oxydans (CGMCC 1788) to improve its water solubility and pharmacokinetic properties. Appl Microbiol Biotechnol 81:47–57CrossRefGoogle Scholar
  15. Kalashnikova GK, Romanova AS, Shchavlinskii AN (1985) Derivatives of the anthracene of Cassia acutifolia Del roots. Khim Farm Zh 19(5):569–573Google Scholar
  16. Kim JH, Bong Kim G, Ko JH, Lee Y, Hur HG, Lim Y, Ahn JH (2006) Molecular cloning, expression, and characterization of a flavonoid glycosyltransferase from Arabidopsis thaliana. Plant Sci 170:897–903CrossRefGoogle Scholar
  17. Kim KH, Moon E, Choi SU, Kim SY, Lee KR (2013) Polyphenols from the bark of Rhus verniciflua and their biological evaluation on antitumor and anti-inflammatory activities. Phytochemistry 92:113–121CrossRefPubMedGoogle Scholar
  18. Ko JH, Kim BG, Ahn JH (2006) Glycosylation of flavonoids with a glycosyltransferase from Bacillus cereus. FEMS Microbiol Lett 258:263–268CrossRefGoogle Scholar
  19. Leatherbarrow RJ (1990) Using linear and non-linear regression to fit biochemical data. Trends Biochem Sci 15:455–458CrossRefPubMedGoogle Scholar
  20. Mohri K, Watanabe Y, Yoshida Y, Satoh M, Isobe K, Sugimoto N, Tsuda Y (2003) Synthesis of glycosylcurcuminoids. Pharm Bull 51:1268–1272CrossRefGoogle Scholar
  21. Olejniczak S, Potrzebowski MJ (2004) Solid state NMR studies and density functional theory (DFT) calculations of conformers of quercetin. Org Biomol Chem 2:2315–2322CrossRefPubMedGoogle Scholar
  22. Quideau S (2011) Triumph for unnatural synthesis. Nature 474:459–460CrossRefPubMedGoogle Scholar
  23. Römer U, Schrader H, Günther N, Nettelstroth N, Frommer WB, Elling L (2004) Expression, purification and characterization of recombinant sucrose synthase 1 from Solanum tuberosum L. for carbohydrate engineering. J Biotechnol 107:135–149CrossRefPubMedGoogle Scholar
  24. Sang SM, Cheng XF, Zhu NQ, Stark RE, Badmaev V, Ghai G, Rosen RT, Ho CT (2001) Flavonol glycosides and novel iridoid glycoside from the leaves of Morinda citrifolia. J Agric Food Chem 49:4478–4481CrossRefPubMedGoogle Scholar
  25. Taleb H, Morris RK, Withycombe CE, Maddocks SE, Kanekanian AD (2016) Date syrup–derived polyphenols attenuate angiogenic responses and exhibits anti-inflammatory activity mediated by vascular endothelial growth factor and cyclooxygenase-2 expression in endothelial cells. Nutr Res 36:636–647CrossRefPubMedGoogle Scholar
  26. Thibodeaux CJ, Melançon CE, Liu HW (2008) Natural-product sugar biosynthesis and enzymatic glycodiversification. Angew Chem Int Ed 47:9814–9859CrossRefGoogle Scholar
  27. Wang HM, Yang Y, Lin L, Zhou WL, Liu MZ, Cheng KD, Wang W (2016) Engineering Saccharomyces cerevisiae with the deletion of endogenous glucosidases for the production of flavonoid glucosides. Microb Cell Fact 15:134–146CrossRefPubMedPubMedCentralGoogle Scholar
  28. Williams GJ, Gantt RW, Thorson JS (2008) The impact of enzyme engineering upon natural product glycodiversification. Curr Opin Chem Biol 12:56–64Google Scholar
  29. Williams GJ, Yang J, Zhang C, Thorson JS (2010) Recombinant E. coli prototype strains for in vivo glycorandomization. ACS Chem Biol 6:95–100CrossRefPubMedPubMedCentralGoogle Scholar
  30. Wu GC, Johnson SK, Bornman JF, Bennett SJ, Fang ZX (2017) Changes in whole grain polyphenols and antioxidant activity of six sorghum genotypes under different irrigation treatments. Food Chem 214:199–207CrossRefPubMedGoogle Scholar
  31. Zhou M, Thorson JS (2011) Asymmetric enzymatic glycosylation of mitoxantrone. Org Lett 13(10):2786–2788CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2017

Authors and Affiliations

  1. 1.State Key Laboratory of Natural MedicinesChina Pharmaceutical UniversityNanjingChina
  2. 2.Jiangsu Key Laboratory of TCM Evaluation and Translational ResearchChina Pharmaceutical UniversityNanjingChina
  3. 3.Institute of Biotechnology for TCM Research, School of Traditional Chinese MedicineChina Pharmaceutical UniversityNanjingChina

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