Crystal structure of a glycoside hydrolase family 68 β-fructosyltransferase from Beijerinckia indica subsp. indica in complex with fructose. Issue 12 (1st December 2020)

• Record Type: Journal Article
• Authors: Tonozuka, Takashi; Kitamura, Junichi; Nagaya, Mika; Kawai, Reika; Nishikawa, Atsushi; Hirano, Katsuaki; Tamura, Keisuke; Fujii, Tadashi; Tochio, Takumi
• Is Part Of: Bioscience, biotechnology, and biochemistry. Volume 84:Issue 12(2020)
• Page Start: 2508
• Page End: 2520
• Publication Date: 2020-12-01
• Subjects: GH68 -- Beijerinckia indica -- β-fructosyltransferase -- fructooligosaccharide -- protein crystallization; Biotechnology -- Periodicals; Biochemistry -- Periodicals; 660.6
• DOI: 10.1080/09168451.2020.1804317
• Languages: English
• ISSNs: 0916-8451

Abstract

ABSTRACT: An enzyme belonging to glycoside hydrolase family 68 (GH68) from Beijerinckia indica subsp. indica NBRC 3744 was expressed in Escherichia coli . Biochemical characterization showed that the enzyme was identified to be a β-fructosyltransferase (BiBftA). Crystallization of a full-length BiBftA was initially attempted, but no crystals were obtained. We constructed a variant in which 5 residues (Pro199-Gly203) and 13 residues (Leu522-Gln534) in potentially flexible regions were deleted, and we successfully crystallized this variant BiBftA. BiBftA is composed of a five-bladed β-propeller fold as in other GH68 enzymes. The structure of BiBftA in complex with fructose unexpectedly indicated that one β-fructofuranose (β-Fru f ) molecule and one β-fructopyranose molecule bind to the catalytic pocket. The orientation of β-Fru f at subsite −1 is tilted from the orientation observed in most GH68 enzymes, presenting a second structure of a GH68 enzyme in complex with the tilted binding mode of β-Fru f. Graphical Abstract: uf0001 Structure of the GH68 enzyme unexpectedly indicated that one β-fructofuranose and one β-fructopyranose bind to the catalytic pocket.