Glucuronic Acid Derivatives in Enzymatic Biomass Degradation: Synthesis and Evaluation of Enzymatic Activity

An essential tool for biotechnology companies in enzyme development for biomass delignification is the access to well-defined model substrates. A deeper understanding of the enzymes substrate specificity can be used to address and optimize enzyme mixtures towards natural, complex substrates. Hence, the chemically synthesized substrates often outcompete those isolated from natural sources in terms of reproducibility, homogeneity and purity.

The first part of this work was the synthesis of two glucuronoxylan fragments designed as model substrates for xylanases. The synthesis involved the use of thioxyloside building blocks in an iterative, linear glycosylation strategy. Two sidechain glucuronate building blocks were synthesized via a divergent strategy from the same thioethylglucose derivative.

In the second part of the project three alkylaromatic and aromatic esters have been prepared as mimics of lignin-carbohydrate complexes found in lignocellulosic biomass, as model substrates for glucuronoyl esterases (GEs). These esters have been used to characterize a novel GE from Cerrena unicolor (CuGE), produced by Novozymes, to obtain insights into the substrate specificity of the enzymes.

HPLC analysis of the enzymatic reactions led to the determination of kinetic parameters that gave information about both bonding affinity and catalytic efficiency.