Transesterification with CE15 glucuronoyl esterase from Cerrena Unicolor reveals substrate specificity

Abstract Glucuronoyl esterases (GE) belonging to family CE15 catalyse the cleavage of ester-linked lignin-carbohydrate complexes (LCCs) in lignocellulosic biomass which exist between the glucuronoyl substitutions of glucuronoxylan and an alcohol on lignin. It is unknown exactly what the enzymes prefer in terms of the alcohol moiety of the substrate, either in the α-benzyl or in the γ-benzyl position. This relates directly to where the ester-LCCs are located on the lignin molecule, and has consequences for how the enzymes potentially interact with lignin. The purpose of this study is to demonstrate how transesterification reactions with a fungal GE from Cerrena Unicolor (CuGE) can reveal the preferences for the type of alcohol. By providing the enzyme with a donor substrate (the methyl ester of either glucuronate or 4-O-methyl-glucuronate) and either one of two acceptor molecules (benzyl alcohol or 3-phenyl-1-propanol) we show that the enzyme preferentially forms the corresponding γ-ester from the methyl ester of 4-O-methyl-glucuronate and 3-phenyl-1-propanol. The results reveal that the enzyme’s substrate preferences are primarily dictated by the presence of the 4-O-methylation on the glucuronoyl donor, and secondly on the type of alcohol.