The variable structural flexibility of the Bacillus circulans β-galactosidase isoforms determines their unique functionalities
Language English Country United States Media print-electronic
Document type Journal Article
PubMed
39353423
DOI
10.1016/j.str.2024.09.005
PII: S0969-2126(24)00374-5
Knihovny.cz E-resources
- Keywords
- cryo-EM, crystal structure, galactooligosaccharide, galactosidase, structural flexibility, substrate specificity, transgalactosylation,
- MeSH
- Bacillus * enzymology MeSH
- Bacterial Proteins chemistry metabolism MeSH
- beta-Galactosidase * chemistry metabolism genetics MeSH
- Cryoelectron Microscopy * MeSH
- Isoenzymes chemistry metabolism MeSH
- Catalytic Domain * MeSH
- Crystallography, X-Ray MeSH
- Models, Molecular * MeSH
- Oligosaccharides metabolism chemistry MeSH
- Protein Isoforms chemistry metabolism MeSH
- Substrate Specificity MeSH
- Publication type
- Journal Article MeSH
- Names of Substances
- Bacterial Proteins MeSH
- beta-Galactosidase * MeSH
- Isoenzymes MeSH
- Oligosaccharides MeSH
- Protein Isoforms MeSH
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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