Fungal feruloyl esterases can catalyze release of diferulic acids from complex arabinoxylan

Feruloyl esterases (FAEs, EC 3.1.1.73) catalyze the hydrolytic cleavage of ester bonds between feruloyl and arabinosyl moieties in arabinoxylans. Recently, we discovered that two bacterial FAEs could catalyze release of diferulic acids (diFAs) from highly substituted, cross-linked corn bran arabinoxylan. Here, we show that several fungal FAEs, notably AnFae1 (Aspergillus niger), AoFae1 (A. oryzae), and MgFae1 (Magnaporthe oryzae (also known as M. grisae)) also catalyze liberation of diFAs from complex arabinoxylan. By comparing the enzyme kinetics of diFA release to feruloyl esterase activity of the enzymes on methyl-and arabinosyl-ferulate substrates we demonstrate that the diFA release activity cannot be predicted from the activity of the enzymes on these synthetic substrates. A detailed structure-function analysis, based on AlphaFold2 modeled enzyme structures and docking with the relevant di-feruloyl ligands, reveal how distinct differences in the active site topology and surroundings may explain the diFA releasing action of the enzymes. Interestingly, the analysis also unveils that the carbohydrate binding module of the MgFae1 may play a key role in the diFA releasing ability of this enzyme. The findings contribute further understanding of the function of FAEs in the deconstruction of complex arabi-noxylans and provide new opportunities for enzyme assisted upgrading of complex bran arabinoxylans.

Automated summary

The study found that esterases from various bacteria and fungi can catalyze the release of diferulic acids from complex arabinoxylans. By comparing the activity of the enzymes on synthetic substrates and diFA release, a detailed structure-function analysis is needed to explain the diFA releasing ability of the enzymes. The findings contribute to a better understanding of the role of esterases in the degradation of complex arabinoxylans and provide new opportunities for enzyme-assisted upgrading of complex bran arabinoxylans.