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Updated: Jun 11, 2025

The Use of a β-lactamase-based Conductimetric Biosensor Assay to Detect Biomolecular Interactions
Published on: February 1, 2018
The variable structural flexibility of the Bacillus circulans β-galactosidase isoforms determines their unique
Michaela Hovorková1, Barbora Kaščáková2, Lucie Petrásková3
1Laboratory of Biotransformation, Institute of Microbiology of the Czech Academy of Sciences, Vídeňská 1083, CZ-14200 Praha4, Czech Republic; Department of Genetics and Microbiology, Faculty of Science, Charles University in Prague, Viničná 5, CZ-12843 Praha2, Czech Republic.
Abstract:
β-Galactosidase from Bacillus circulans ATCC 31382 (BgaD) is a biotechnologically important enzyme for the synthesis of β-galactooligosaccharides (GOS). Among its four isoforms, isoform A (BgaD-A) has distinct synthetic properties. Here, we present cryoelectron microscopy (cryo-EM) structures of BgaD-A and compare them with the known X-ray crystal structure of isoform D (BgaD-D), revealing substantial structural divergences between the two isoforms. In contrast to BgaD-D, BgaD-A features a flexible Big-4 domain and another enigmatic domain. The newly identified flexible region in BgaD-A is termed as "barrier domain 8," and serves as a barricade, obstructing the access of longer oligosaccharide substrates into the active site of BgaD-A. The transgalactosylation reactions catalyzed by both isoforms revealed that BgaD-A has a higher selectivity than BgaD-D in the earlier stages of the reaction and is prevailingly directed to shorter galactooligosaccharides. This study improves our understanding of the structural determinants governing β-galactosidase catalysis, with implications for tailored GOS production.
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