Functional and structural diversity in GH62 α-L-arabinofuranosidases from the thermophilic fungus Scytalidium thermophilum
Abstract
The genome of the thermophilic fungus Scytalidium thermophilum (strain CBS 625.91) harbours a wide range of genes involved in carbohydrate degradation, including three genes, abf62A, abf62B and abf62C, predicted to encode glycoside hydrolase family 62 (GH62) enzymes. Transcriptome analysis showed that only abf62A and abf62C are actively expressed during growth on diverse substrates including straws from barley, alfalfa, triticale and canola. The abf62A and abf62C genes were expressed in Escherichia coli and the resulting recombinant proteins were characterized. Calcium-free crystal structures of Abf62C in apo and xylotriose bound forms were determined to 1.23 and 1.48 Å resolution respectively. Site-directed mutagenesis confirmed Asp55, Asp171 and Glu230 as catalytic triad residues, and revealed the critical role of non-catalytic residues Asp194, Trp229 and Tyr338 in positioning the scissile α-L-arabinofuranoside bond at the catalytic site. Further, the +2R substrate-binding site residues Tyr168 and Asn339, as well as the +2NR residue Tyr226, are involved in accommodating long-chain xylan polymers. Overall, our structural and functional analysis highlights characteristic differences between Abf62A and Abf62C, which represent divergent subgroups in the GH62 family.
- Authors:
-
- Univ. of Toronto, ON (Canada). Dept. of Chemical Engineering and Applied Chemistry.
- Argonne National Lab. (ANL), Argonne, IL (United States). Structureal Biology Center.
- Concordia Univ., Montreal, Quebec (Canada). Centre for Structural and Functional Genomics.
- Concordia Univ., Montreal, Quebec (Canada). Centre for Structural and Functional Genomics and Dept. of Chemistry and Biochemistry.
- Concordia Univ., Montreal, Quebec (Canada). Centre for Structural and Functional Genomics and Dept. of Biology.
- Univ. of Toronto, ON (Canada); Dept. of Chemical Engineering and Applied Chemistry.
- Publication Date:
- Research Org.:
- Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
- Sponsoring Org.:
- USDOE Office of Science (SC), Biological and Environmental Research (BER)
- OSTI Identifier:
- 1212712
- Grant/Contract Number:
- AC02-06CH11357
- Resource Type:
- Journal Article: Accepted Manuscript
- Journal Name:
- Microbial Biotechnology (Online)
- Additional Journal Information:
- Journal Volume: 8; Journal Issue: 3; Journal ID: ISSN 1751-7915
- Publisher:
- Wiley
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 60 APPLIED LIFE SCIENCES
Citation Formats
Kaur, Amrit Pal, Nocek, Boguslaw P., Xu, Xiaohui, Lowden, Michael J., Leyva, Juan Francisco, Stogios, Peter J., Cui, Hong, Leo, Rosa Di, Powlowski, Justin, Tsang, Adrian, and Savchenko, Alexei. Functional and structural diversity in GH62 α-L-arabinofuranosidases from the thermophilic fungus Scytalidium thermophilum. United States: N. p., 2015.
Web. doi:10.1111/1751-7915.12168.
Kaur, Amrit Pal, Nocek, Boguslaw P., Xu, Xiaohui, Lowden, Michael J., Leyva, Juan Francisco, Stogios, Peter J., Cui, Hong, Leo, Rosa Di, Powlowski, Justin, Tsang, Adrian, & Savchenko, Alexei. Functional and structural diversity in GH62 α-L-arabinofuranosidases from the thermophilic fungus Scytalidium thermophilum. United States. https://doi.org/10.1111/1751-7915.12168
Kaur, Amrit Pal, Nocek, Boguslaw P., Xu, Xiaohui, Lowden, Michael J., Leyva, Juan Francisco, Stogios, Peter J., Cui, Hong, Leo, Rosa Di, Powlowski, Justin, Tsang, Adrian, and Savchenko, Alexei. 2015.
"Functional and structural diversity in GH62 α-L-arabinofuranosidases from the thermophilic fungus Scytalidium thermophilum". United States. https://doi.org/10.1111/1751-7915.12168. https://www.osti.gov/servlets/purl/1212712.
@article{osti_1212712,
title = {Functional and structural diversity in GH62 α-L-arabinofuranosidases from the thermophilic fungus Scytalidium thermophilum},
author = {Kaur, Amrit Pal and Nocek, Boguslaw P. and Xu, Xiaohui and Lowden, Michael J. and Leyva, Juan Francisco and Stogios, Peter J. and Cui, Hong and Leo, Rosa Di and Powlowski, Justin and Tsang, Adrian and Savchenko, Alexei},
abstractNote = {The genome of the thermophilic fungus Scytalidium thermophilum (strain CBS 625.91) harbours a wide range of genes involved in carbohydrate degradation, including three genes, abf62A, abf62B and abf62C, predicted to encode glycoside hydrolase family 62 (GH62) enzymes. Transcriptome analysis showed that only abf62A and abf62C are actively expressed during growth on diverse substrates including straws from barley, alfalfa, triticale and canola. The abf62A and abf62C genes were expressed in Escherichia coli and the resulting recombinant proteins were characterized. Calcium-free crystal structures of Abf62C in apo and xylotriose bound forms were determined to 1.23 and 1.48 Å resolution respectively. Site-directed mutagenesis confirmed Asp55, Asp171 and Glu230 as catalytic triad residues, and revealed the critical role of non-catalytic residues Asp194, Trp229 and Tyr338 in positioning the scissile α-L-arabinofuranoside bond at the catalytic site. Further, the +2R substrate-binding site residues Tyr168 and Asn339, as well as the +2NR residue Tyr226, are involved in accommodating long-chain xylan polymers. Overall, our structural and functional analysis highlights characteristic differences between Abf62A and Abf62C, which represent divergent subgroups in the GH62 family.},
doi = {10.1111/1751-7915.12168},
url = {https://www.osti.gov/biblio/1212712},
journal = {Microbial Biotechnology (Online)},
issn = {1751-7915},
number = 3,
volume = 8,
place = {United States},
year = {Fri May 01 00:00:00 EDT 2015},
month = {Fri May 01 00:00:00 EDT 2015}
}
Web of Science
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