title: Enzymatic synthesis of glycosylated puerarin using maltogenic amylase from Bacillus stearothermophilus expressed in Bacillus subtilis.
abstract: BACKGROUND
  The maltogenic amylase from Bacillus stearothermophilus (BSMA) is a valuable biocatalyst that has been used to transglycosylate natural glycosides to improve solubility. To ensure safety, BSMA was produced in Bacillus subtilis, using new shuttle vector-based expression vectors. The transglycosylation of puerarin was also conducted with crude BSMA and analyzed.
  
  
  RESULTS
  Two expression systems, each containing one of the promoters from the genes encoding Bacillus licheniformis maltogenic amylase (BLMA) and an alpha-amylase from B. subtilis NA64 (amyR2), were constructed. The amyR2 promoter system was chosen as the best system; it yielded 107 mg of pure BSMA from a 2 L culture. In the transglycosylation reactions of puerarin using crude BSMA, relative amounts for maltosyl-alpha-(1 --> 6)-puerarin, glucosyl-alpha-(1 --> 6)-puerarin, glucosyl-alpha-(1 --> 3)-puerarin, and puerarin were determined as 26:18:7:49. A two-step purification process, including gel permeation chromatography, yielded 1.7 g of the transfer products from 3 g of puerarin.
  
  
  CONCLUSIONS
  The crude BSMA produced from a host generally recognized as safe (B. subtilis) can be used to transglycosylate various functional compounds. The expression system developed in this study will be helpful for the production of other food-grade enzymes by B. subtilis.
authors.affiliation: Center for Agricultural Biomaterials and Department of Food Science and Biotechnology, Seoul National University, Seoul 151-921, Korea.
authors.forename: Chung-Hyo
authors.initials: CH
authors.lastname: Choi
authors.forename: Seung-Hee
authors.initials: SH
authors.lastname: Kim
authors.forename: Jun-Hyuck
authors.initials: JH
authors.lastname: Jang
authors.forename: Jong-Tae
authors.initials: JT
authors.lastname: Park
authors.forename: Jae-Hoon
authors.initials: JH
authors.lastname: Shim
authors.forename: Young-Wan
authors.initials: YW
authors.lastname: Kim
authors.forename: Kwan-Hwa
authors.initials: KH
authors.lastname: Park
mesh_terms: D000681:Amylases / Q000235:genetics / Q000378:metabolism*; D001412:Bacillus subtilis / Q000201:enzymology* / Q000235:genetics; D004274:DNA, Recombinant; D015870:Gene Expression; D005798:Genes, Bacterial; D005818:Genetic Engineering / Q000379:methods; D001411:Geobacillus stearothermophilus / Q000201:enzymology* / Q000235:genetics; D006031:Glycosylation; D007529:Isoflavones / Q000096:biosynthesis*; D010957:Plasmids; D011401:Promoter Regions, Genetic; D029908:Pueraria / Q000737:chemistry; D012995:Solubility
chemical_list: D004274:DNA, Recombinant; D007529:Isoflavones; D000681:Amylases; C033607:puerarin