Photocrosslinking approaches are an important strategy to covalently capture biomolecular interactions. Covalent capture by photocrosslinking is particularly powerful for glycan-dependent interactions because these binding events are typically low-affinity and unable to withstand common purification protocols. Implementation of photocrosslinking methods is based on the incorporation of a photoactivatable crosslinking functional group onto glycan-containing molecules. Upon activation with UV radiation, the photocrosslinking group is converted to a reactive species, which forms a covalent bond with neighboring molecules. Resulting crosslinked complexes can be analyzed by immunological and mass spectrometry techniques to identify glycan-dependent binding partners. In some cases, photocrosslinker incorporation can be achieved in living cells, enabling detection of glycan-dependent interactions in their native context. In this mini-review, we describe new sugar-based photocrosslinking reagents and illustrate how they have been used to photochemically characterize glycan-dependent interactions.
The Bifidobacterium genus harbours several health promoting members of the gut microbiota, which display metabolic specialization by preferentially utilizing dietary or host-derived β-galactosides. To approach a deeper understanding of the β-galactoside metabolism in the gut niche, the overall study investigated the bioinformatics, biochemistry and structures of glycoside hydrolase family 42 (GH42) β-galactosidases from Bifidobacterium species associated with the human gut. GH42 β-galactosidases display a large variety of sub-specificities in accordance with the diversity and complexity of β-galactosides available in the gut. The variety of sub-specificities is evident in a phylogenetic distribution where GH42 β-galactosidases segregate according to function. The first function-structure insight in GH42 illustrates that diversity manifested in sub-specificities, correlates through subtle changes in loop regions in the near vicinity of the site of catalysis. The occurrence of, multiple GH42 β-galactosidases with diverse sub-specificities within a single Bifidobacterium strain and the distinct differences among all of them from various species, emphasizes the importance and diversity of β-galactoside metabolism in bifidobacteria.