Functional expression and characterization of a glucuronoyl esterase from the fungus Neurospora crassa: identification of novel consensus sequences containing the catalytic triad

The Journal of General and Applied Microbiology

Manabu Arioka¹  Hung Hiep Huynh¹ 

[1] Department of Biotechnology, The University of Tokyo

关键词: carbohydrate esterase family 15;    glucuronoyl esterase;    hemicellulose;    lignin-carbohydrate complex;    Neurospora crassa;   

DOI  :  10.2323/jgam.2016.03.004

学科分类：微生物学和免疫学

来源: Applied Microbiology, Molecular and Cellulrar Biosciences Research Foundation

【 摘 要 】

The complete hydrolysis of lignocellulose requires the actions of a variety of enzymes, including those that cleave the linkage between lignin and hemicellulose. The enzyme glucuronoyl esterase (GE) that constitutes a novel family of carbohydrate esterases, CE15, has been shown to display a unique ability to cleave the ester linkage between lignin alcohols and xylan-bound 4-O-methyl-D-glucuronic acid of hemicellulose. We herein report identification, expression, and functional characterization of a new GE, NcGE, from the filamentous fungus Neurospora crassa. C-terminally c-myc and hexahistidine-tagged NcGE was heterologously expressed in the methylotrophic yeast Pichia pastoris. NcGE purified from the culture supernatant through Ni-NTA and anion exchange chromatographies showed the ability to hydrolyze the substrate 3-(4-methoxyphenyl) propyl methyl 4-O-methyl-α-D-glucopyranosiduronate, which mimics the ester linkage of 4-O-methyl-D-glucuronic acid in lignin-carbohydrate complexes (LCCs). This esterase showed the characteristic of a mesophilic enzyme with the temperature optimum at 40–50°C, and displayed the optimal activity at pH 7 and broad pH stability. Based on the alignment of NcGE with other GEs so far characterized, we propose novel consensus sequences for GEs containing the catalytic triad.

【 授权许可】

Unknown