Enzymatic transfucosylation for synthesis of human milk oligosaccharides

Human milk oligosaccharides (HMOs) are a family of structurally distinct carbohydrate oligomers present in human milk. HMOs protect breastfed babies against infection and promote the development of infant health and cognition. In the gut, fucosylated HMOs in particular function as decoy receptors that intercept epithelial attachment of enteric pathogens and hence help reduce infection. Infant formulae made from bovine milk are essentially devoid of HMOs, which creates a large impetus for biosynthetic production of HMOs. Certain microbial α-L-fucosidases (EC 3.2.1.51, EC 3.2.1.111), specifically various retaining α-L-fucosidases of glycoside hydrolase family 29 (GH29), are capable of catalysing transfucosylation. The use of GH29 α-L-fucosidases to promote transfucosylation reactions thus represents a strategy for biocatalytic synthesis of fucosylated HMOs. The purpose of this review is to present the current knowledge on the use of such α-L-fucosidases for synthesis of fucosylated HMOs by enzymatic transfucosylation. We summarize the available data obtained for both wild type and engineered microbial α-L-fucosidases, discuss enzyme and substrate sources, and review factors governing transglycosylation performance, particularly the use of protein engineering. We describe the mechanistic reaction details of α-l-fucosidase transfucosylation, and examine details of enzyme mutation strategies promoting transfucosylation. Finally, we list recommendations for future reaction targets based on currently abundant substrate sources.