Premium
Glycosylation of Ganoderic Acid A via Recombinant Glycosyltransferase of Bacillus subtilis Under Acidic Operating Condition
Author(s) -
Wu Jiumn-Yih,
Chang Te-Sheng,
Chiang Chien-Min,
Wang Tzi-Yuan
Publication year - 2020
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2020.34.s1.03156
Subject(s) - bacillus subtilis , glycosylation , glycosyltransferase , chemistry , biochemistry , trisaccharide , escherichia coli , bacillus licheniformis , enzyme , nucleotide sugar , uridine diphosphate glucose , glycoside , biotransformation , uridine diphosphate , bacteria , stereochemistry , biology , gene , genetics
Ganoderic acid A (GAA) is a triterpenoid with identified effective activity, isolated from Ganoderma lucidum . Glycosylation of triterpenoid might improve its anti‐toxin bioactivity and also increase both water solubility and physical‐chemical stability. Glycosylation is carried out by glycosyltransferases (GT, EC 2.4.x.y), a type of enzyme that utilize a nucleotide‐activated sugar donor, such as uridine diphosphate (UDP)‐glucose, to transfer the sugar portion to a sugar‐linked molecule. We have utilized Bacillus sp. GA A07, isolated from intestinal bacteria of Zebrafish, to biotransform GAA to GAA‐15‐ O ‐β‐glucoside 1. Our previous study also showed that the Bacillus subtilis , ATCC 6633 strain, could biotransform GAA to compound, GAA‐15‐O‐b‐glucoside, and one minor compound in Fig. 1. We selected five GT genes‐BsGT110, BsGT292, BsGT296, BsGT398, and BsGT489 from the used strains, and successfully overexpressed and cloned them in Escherichia coli . Two glycosyltransferases (BsGT398 and BsGT489) have shown the capability to catalyze GAA to GAA‐15‐O‐β‐glucoside under some specific conditions. However, the chemical structure of minor compound 2 and its corresponding enzyme remain elusive. In the present study, we identified BsGT110, a GT from the used B. subtilis strain, for the biotransformation of GAA into this compound under acidic glycosylation. BsGT110 showed an optimal glycosylation activity toward GAA at pH 6 but lost most of its activity at pH 8. Through a designed operating production, this product 2 was successfully isolated using preparative high‐performance liquid chromatography and was identified to be a new triterpenoid glucoside (GAA‐26‐O‐β‐glucoside) by mass and nuclear magnetic resonance spectroscopy. We identified that BsGT110 is an unique GT that have a specific activity on the glycosylation of triterpenoid at the C‐26 position under acidic conditions (in Fig. 2) but lost most of its activity at alkaline ones, suggesting that acidic solutions may enhance the catalysis activity of this and similar types of GTs toward triterpenoids. Support or Funding Information This research was financially supported by the grants of the Ministry of Science and Technology, Taiwan (Project No. MOST 108‐2221‐E‐507 ‐008 ‐ and MOST 108‐2221‐E‐024 ‐008 ‐MY2).UPLC analysis of the biotranformation of GAA by BsGT110Glycosylation of GAA by BsGT110 under the acidic conditionReference 1 Te-Sheng Chang , Chien-Min Chiang , Tzi-Yuan Wang , Chun-Hsien Lee , Yu-Wen Lee , and Jiumn-Yih Wu * , “ New Triterpenoid from Novel Triterpenoid 15-O-Glycosylation on Ganoderic Acid A by Intestinal Bacteria of Zebrafish ”, Molecules , 2018 , 23 , 2345 – 2354 . 2 Te-Sheng Chang , Jiumn-Yih Wu , Tzi-Yuan Wang , Kun-Yuan Wu and Chien-Min Chiang * , “ Uridine Diphosphate-Dependent Glycosyltransferases from Bacillus subtilis ATCC 6633 Catalyze 15-O-Glycosylation of Ganoderic Acid A ”, International Journal of Molecular Sciences , 2018 , 19 , 3469 – 3480