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Change of Bacillus cereus Flavonoid O-Triglucosyltransferase into Flavonoid O-Monoglucosyltransferase by error-prone polymerase chain reaction
1Department of Bioscience and Biotechnology, Bio/Molecular Informatics Center, Konkuk University, Seoul 143-701, Korea, 2National Institute of Animal Science, Rural Development Administration, Suwon 441-706, Korea
J. Microbiol. Biotechnol. 2010; 20(10): 1393-1396
Published October 28, 2010 https://doi.org/10.4014/jmb.1003.03005
Copyright © The Korean Society for Microbiology and Biotechnology.
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J. Microbiol. Biotechnol. 2010; 20(10): 1393-1396
Published online October 28, 2010 https://doi.org/10.4014/jmb.1003.03005
Copyright © The Korean Society for Microbiology and Biotechnology.
Change of Bacillus cereus Flavonoid O-Triglucosyltransferase into Flavonoid O-Monoglucosyltransferase by error-prone polymerase chain reaction
Na Ri Jung 1, Enu Ji Joe 1, Bong-Gyu Kim 1, Byoung Chan Ahn 1, Jun Cheol Park 2, Youhoon Chong 1 and Joong-Hoon Ahn 1*
1Department of Bioscience and Biotechnology, Bio/Molecular Informatics Center, Konkuk University, Seoul 143-701, Korea, 2National Institute of Animal Science, Rural Development Administration, Suwon 441-706, Korea
Abstract
The attachment of sugar to flavonoids enhances their solubility. Glycosylation is performed primarily by uridine diphosphate-dependent glycosyltransferases (UGTs). The UGT from Bacillus cereus, BcGT-1 transferred three glucose molecules into kaempferol. The structural analysis of BcGT-1 showed that its substrate binding site is wider than that of flavonoid monoglucosyltransferase of plant. In order to create monoglucosyltransferase from BcGT-1, error-prone polymerase chain reaction (PCR) was performed. We analyzed 150 clones. Among them, two mutants generated only kaempferol O-monoglucoside, albeit with reduced reactivity. Unexpectedly, the two mutants harbored mutations in the amino acids located outside of the active sites. Based on the modeled structure of BcGT-1, it was proposed that the local change in the secondary structure of BcGT-1 caused the alteration of triglucosyltransferase into monoglucosyltransferase.
Keywords: Bacillus cereus, Flavonoid, Glycosyltransferase