Unexpected Anomeric Acceptor Preference Observed Using dDNP NMR for Transglycosylation Studies of β‑Galactosidases

• Published in: Biochemistry (Easton) Vol. 59; no. 31; pp. 2903 - 2908
• Main Authors: Kjeldsen, Christian, Ardenkjær-Larsen, Jan H, Duus, Jens. Ø
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 11.08.2020
• Subjects: beta-Galactosidase - metabolism; Galactose - chemistry; Galactose - metabolism; Glucose - chemistry; Glucose - metabolism; Glycosylation; Kinetics; Nuclear Magnetic Resonance, Biomolecular; Stereoisomerism; Substrate Specificity
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/acs.biochem.0c00390

The transglycosylation abilities of β-galactosidases were investigated using hyperpolarized [U–13C,U–2H]­glucose as an acceptor and o-nitrophenyl β-galactopyranoside as a donor. Several products were readily observable, and at least in the case when O3 acted as an acceptor, the enzymes showed a clear selectivity toward the β-anomer of glucose. Additionally, it was possible to determine the relative hydrolysis rates of the formed transglycosylation products, providing information on the selectivity as well. Using this method, the transglycosylation abilities of the enzymes could be studied at a very high temporal resolution as well as with high sensitivity, and due to the relative ease of the setup, this method could be more generally applied to investigate glycosidases.