Expanding the feruloyl esterase gene family of Aspergillus niger by characterization of a feruloyl esterase, FaeC

Abstract

A feruloyl esterase (FAE) from Aspergillus niger N402, FaeC was heterologously produced in Pichia pastoris X-33 in a yield of 10mg/L. FaeC was most active at pH 7.0 and 50°C, and showed broad substrate specificity and catalyzed the hydrolysis of methyl 3,4-dimethoxycinnamate, ethyl ferulate, methyl ferulate, methyl p-coumarate, ethyl coumarate, methyl sinapate, and methyl caffeate. The enzyme released both ferulic acid and p-coumaric acid from wheat arabinoxylan and sugar beet pectin (up to 3mg/g polysaccharide), and acted synergistically with a commercial xylanase increasing the release of ferulic acid up to six-fold. The expression of faeC increased over time in the presence of feruloylated polysaccharides. Cinnamic, syringic, caffeic, vanillic and ferulic acid induced the expression of faeC. Overall expression of faeC was very low in all tested conditions, compared to two other A. niger FAE encoding genes, faeA and faeB. Our data showed that the fae genes responded differently towards the feruloylated polysaccharides and tested monomeric phenolic compounds suggesting that the corresponding FAE isoenzymes may target different substrates in a complementary manner. This may increase the efficiency of the degradation of diverse plant biomass.

Evolutionary adaptation of Aspergillus niger for increased ferulic acid tolerance

“…Both induced LPMOs have been predicted to act on cellulose (Borin et al ) and one of them (NRRL3_3929) has been suggested to be regulated by AraR (Gruben et al ). The transcriptional profile of the FAE encoding genes correlated with previous observations (de Vries et al ; Dilokpimol et al ). Ferulic acid can be linked to l ‐arabinose in xylan, which could explain why the other xylanolytic genes were also induced by ferulic acid.…”

The Synthetic Potential of Fungal Feruloyl Esterases: A Correlation with Current Classification Systems and Predicted Structural Properties

“…P. pastoris transformants containing FAEs from Aspergillus species, Ceriporiopsis subvermispora, and Galerina marginata were grown according to previous reports [26,28,30,32]. Induction was done for 96 h at 28 • C with methanol being supplemented to 0.5% (v/v) every 24 h. Culture supernatants were harvested (4000× g, 4 • C, 20 min), filtered (0.22 µm; Merck Millipore, Darmstadt, Germany), concentrated (10 kDa cut off; Merck Millipore, Germany), and stored at −20 • C prior to further analysis.…”

Characterization of a feruloyl esterase from Aspergillus terreus facilitates the division of fungal enzymes from Carbohydrate Esterase family 1 of the carbohydrate‐active enzymes (CAZy) database

“…As an example case to confirm the consistency of the substrate specificity within the subfamilies of CE1, we heterologously expressed and characterized A. terreus FaeD. This enzyme is a member of subfamily 5 (Figs and S1), which also contains the recently characterized AnFaeC from A. niger (Dilokpimol et al ., ) in a different branch. The Aspergilli appear to have abundant numbers of putative FAE‐encoding genes, and previously clear differences have been reported between the three characterized FAEs from A. niger (de Vries et al ., ; Dilokpimol et al ., ).…”

“…These include N. crassa FaeD (Crepin et al ., ), A. sydowii FaeC and FaeD1 (Dilokpimol et al ., ), A. nidulans FaeC (Debeire et al ., ) and A. niger FaeC (Dilokpimol et al ., ), as well as AtFaeD characterized in this study, thus supporting that CE1 subfamily 5 is also a FAE subfamily. In addition to the artificial FAE substrates, several of the functionally characterized subfamily 5 enzymes have also been shown to release ferulic acid from natural plant biomass substrates, such as wheat arabinoxylan (WAX) and SBP (Crepin et al ., ; Dilokpimol et al ., ), or hydrolyse feruloylated oligosaccharides (Debeire et al ., ).…”