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Title: Novel Family of Carbohydrate Esterases, Based on Identification of the Hypocrea jecorina Acetyl Esterase Gene

Author
item LI, XIN-LIANG - TACOMA, WA
item Skory, Christopher - Chris
item Cotta, Michael
item PUCHART, VLADIMIR - SLOVAK ACADEMY
item BIELY, PETER - SLOVAK ACADEMY

Submitted to: Applied and Environmental Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/7/2008
Publication Date: 11/1/2008
Citation: Li, X., Skory, C.D., Cotta, M.A., Puchart, V., Biely, P. 2008. Novel family of carbohydrate esterases, based on identification of the Hypocrea jecorina Acetyl Esterase Gene. Applied and Environmental Microbiology. 74(24):7482-7489.

Interpretive Summary: Energy crops and crop residues are abundant sources of plant fiber (lignocellulose) that could be used to produce a variety of bioproducts. However, use of these agricultural biomass resources to produce fuels and chemicals will require the development of efficient enzyme systems in order to convert the complex carbohydrates contained within plant cell walls to fermentable sugars. Carbohydrate esterases are a group of such enzymes that help with the conversion of the hemicellulose portion of plant cell walls. The current research identified and characterized an acetyl esterase from the commercially important fungus, Hypocrea jecorina, and found that this and perhaps related enzymes constitute a new family of esterases that can be used for lignocellulose bioconversion. This information will help in the development of biochemical conversion process for the conversion of plant fiber to fuels and chemicals.

Technical Abstract: Plant cell walls have been shown to contain acetyl groups in hemicelluloses and pectin. The gene, ae1, encoding the acetyl esterase (Ae1) of Hypocrea jecorina was identified by amino terminal sequencing, peptide mass spectrometry, and genomic sequence analyses. The coded polypeptide had 348 amino acid residues with the first 19 serving as a secretion signal peptide. The calculated molecular mass and isoelectric point of the secreted enzyme were 37088 Da and pH 5.89, respectively. No significant homology was found between the predicated Ae1 protein and carbohydrate esterases of known families, but putative ae1 orthologs were found in genomes of many fungi and bacteria that produce cell wall-degrading enzymes. The ael transcript levels were high when the fungal cells were induced with sophorose, cellulose, oat spelt xylan, lactose, and arabinose. The recombinant Ae1, produced by H. jecorina transformed with Ae1 under the cellobiohydrolase I promoter, displayed properties similar to those reported for the native enzyme. The enzyme hydrolyzed acetate ester bond specifically. Using 4-nitrophenyl acetate as substrate, the activity of the recombinant enzyme was enhanced by D-xylose, D-glucose, cellobiose, D-galactose, and xylooligosaccharides, but not by arabinose, mannose, or lactose. Using 4-nitrophenyl beta-D-xylopyranoside monoacetate as substrate in a beta-xylosidase coupled assay, Ae1 hydrolyzed positions 3 and 4 with the same efficiency, while the H. jecorina acetylxylan esterase exclusively deacetylated the position 2 acetyl group. The data presented demonstrated that Ae1 and perhaps other homologous microbial proteins represent a new family of esterases for lignocellulose biodegradation.