Characterization and alteration of product specificity of Beijerinckia indica subsp. indica β-fructosyltransferase

Ding Li, Yuki Miyasaka, Arisa Kubota, Takuma Kozono, Yoshikazu Kitano, Nobumitsu Sasaki, Tadashi Fujii, Takumi Tochio, Yoshihiro Kadota, Atsushi Nishikawa, Takashi Tonozuka

Research output: Contribution to journalArticlepeer-review

1 Citation (Scopus)

Abstract

The trisaccharide 1-kestose, a major constituent of fructooligosaccharide, has strong prebiotic effects. We used high-performance liquid chromatography and 1H nuclear magnetic resonance spectroscopy to show that BiBftA, a β-fructosyltransferase belonging to glycoside hydrolase family 68, from Beijerinckia indica subsp. indica catalyzes transfructosylation of sucrose to produce mostly 1-kestose and levan polysaccharides. We substituted His395 and Phe473 in BiBftA with Arg and Tyr, respectively, and analyzed the reactions of the mutant enzymes with 180 g/L sucrose. The ratio of the molar concentrations of glucose and 1-kestose in the reaction mixture with wild-type BiBftA was 100:8.1, whereas that in the reaction mixture with the variant H395R/F473Y was 100:45.5, indicating that H395R/F473Y predominantly accumulated 1-kestose from sucrose. The X-ray crystal structure of H395R/F473Y suggests that its catalytic pocket is unfavorable for binding of sucrose while favorable for transfructosylation.

Original languageEnglish
Pages (from-to)981-990
Number of pages10
JournalBioscience, Biotechnology and Biochemistry
Volume87
Issue number9
DOIs
Publication statusPublished - 01-09-2023

All Science Journal Classification (ASJC) codes

  • General Medicine

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