Unnatural C6‐Substituted d ‐Galactosides as Attenuated Substrates of Glycosyl Hydrolases

The covalent attachment of pendant groups to the carbons of d ‐pyranosides can influence their ability to be recognized by proteins that bind natural sugars. These carbon‐modified sugars have received only limited attention, and systematic investigations on their ability to act as substrates of glycosyl hydrolases have not been reported. Here, C6‐substituted β‐ d ‐galactosides are synthesized and evaluated as putative substrates of β‐glycosyl hydrolases. The results indicated that the size of the group at C6—methyl vs. di‐methyl vs. phenyl—attenuated their ability to act as substrates. The C6‐methyl derivative was a weak substrate, but the C6‐dimethyl and C6‐phenyl compounds were not. The new β‐galactosides can be leveraged as tool compounds in the search for glycosidases with novel activity and as starting points for enzyme‐engineering campaigns.