Cloning, expression of a feruloyl esterase from Aspergillus usamii E001 and its applicability in generating ferulic acid from wheat bran

Abstract

A cDNA gene (AufaeA), which encodes a mature polypeptide of the type-A feruloyl esterase from Aspergillus usamii E001 (abbreviated to AuFaeA), was cloned and heterologously expressed in Pichia pastoris GS115. One transformant, labeled as P. pastoris GSFaeA4-8, expressing the highest recombinant AuFaeA (reAuFaeA) activity of 10.76 U/ml was selected by the flask expression test. The expressed reAuFaeA was purified to homogeneity with an apparent molecular weight of 36.0 kDa by SDS-PAGE analysis, and characterized using the model substrate of methyl ferulate (MFA). The purified reAuFaeA was optimally active at pH 5.0 and 45 °C, and highly stable at pH 4.0-6.5 and 45 °C or below. Its activity was not significantly affected by metal ions tested and EDTA. The K m and V max of reAuFaeA towards MFA were 4.64 mM and 115.5 U/mg, respectively. High-performance liquid chromatography analysis showed that only 9.7 % of total alkali-extractable ferulic acid (FA) was released from destarched wheat bran by reAuFaeA alone. The released FA increased to 36.5 % when reAuFaeA was used together with a recombinant Aspergillus usamii GH family 11 xylanase A, indicating a synergistic interaction between them.

Enrichment of maize and triticale bran with recombinant Aspergillus tubingensis ferulic acid esterase

“…Anion exchange chromatography enabled 7-fold purification and a 48.2% recovery of FAE activity (Table 1). The specific activity of the purified AtFAEA on 4NFP as substrate was 77.6 U/mg protein, which was higher than recombinantly expressed feruloyl esterases from Aspergillus usamii (44.9 U/mg) (Gong et al 2013), A. nidulans (21.7 U/mg) (Debeire et al 2012), A. awamori (9.01 U/mg) (Koseki et al 2005) and A. oryzae (0.57 U/mg) (Koseki et al 2009). The purified AtFAE displayed a 36 kDa protein species on SDS-PAGE (Fig.…”

Thermotolerant hemicellulolytic and cellulolytic enzymes from Eupenicillium parvum 4-14 display high efficiency upon release of ferulic acid from wheat bran

“…; Gong et al . ), and has been identified as a key enzyme in releasing FA from plant materials (Uraji et al . ).…”

Molecular Modeling and MM-PBSA Free Energy Analysis of Endo-1,4-β-Xylanase from Ruminococcus albus 8

“…The main chain of xylan is composed of 1,4- d -xylose subunits, which is usually decorated with various side chain residues of 1,2-α- d -glucuronic acid, or its 4- O -methyl ethers, 1,3-α- l -arabinose, and/or O -acetyl groups in the 2 and 3 positions. Due to structural complexity, several xylanolytic enzymes are required to release the substituents and sugars from the various xylans, including endo-1,4-β-xylanases (EC 3.2.1.8) [7], acetyl xylan esterases (EC 3.1.1.72) [8], feruloyl esterases (EC 3.1.1.73) [9], α- l -arabinofuranosidases (EC 3.2.1.55) [10], α-glucuronidases (EC 3.2.1.139) [11], and β- d -xylosidase (EC 3.2.1.37) [12]. Among these enzymes, endo-1,4-β-xylanases are believed to be the most valuable in industrial applications.…”