Characterization of a highly xylose tolerant β-xylosidase isolated from high temperature horse manure compost
| dc.contributor.author | Ndata, Kanyisa | |
| dc.contributor.author | Nevondo, Walter | |
| dc.contributor.author | Cekuse, Bongi | |
| dc.contributor.author | van Zyl, Leonardo J. | |
| dc.contributor.author | Trindade, Marla | |
| dc.date.accessioned | 2021-11-02T13:40:01Z | |
| dc.date.available | 2021-11-02T13:40:01Z | |
| dc.date.issued | 2021-10-24 | |
| dc.date.updated | 2021-10-31T04:18:30Z | |
| dc.description.abstract | Background There is a continued need for improved enzymes for industry. β-xylosidases are enzymes employed in a variety of industries and although many wild-type and engineered variants have been described, enzymes that are highly tolerant of the products produced by catalysis are not readily available and the fundamental mechanisms of tolerance are not well understood. Results Screening of a metagenomic library constructed of mDNA isolated from horse manure compost for β-xylosidase activity identified 26 positive hits. The fosmid clones were sequenced and bioinformatic analysis performed to identity putative β-xylosidases. Based on the novelty of its amino acid sequence and potential thermostability one enzyme (XylP81) was selected for expression and further characterization. XylP81 belongs to the family 39 β-xylosidases, a comparatively rarely found and characterized GH family. The enzyme displayed biochemical characteristics (KM—5.3 mM; Vmax—122 U/mg; kcat—107; Topt—50 °C; pHopt—6) comparable to previously characterized glycoside hydrolase family 39 (GH39) β-xylosidases and despite nucleotide identity to thermophilic species, the enzyme displayed only moderate thermostability with a half-life of 32 min at 60 °C. Apart from acting on substrates predicted for β-xylosidase (xylobiose and 4-nitrophenyl-β-D-xylopyranoside) the enzyme also displayed measurable α-L-arabainofuranosidase, β-galactosidase and β-glucosidase activity. A remarkable feature of this enzyme is its ability to tolerate high concentrations of xylose with a Ki of 1.33 M, a feature that is highly desirable for commercial applications. Conclusions Here we describe a novel β-xylosidase from a poorly studied glycosyl hydrolase family (GH39) which despite having overall kinetic properties similar to other bacterial GH39 β-xylosidases, displays unusually high product tolerance. This trait is shared with only one other member of the GH39 family, the recently described β-xylosidases from Dictyoglomus thermophilum. This feature should allow its use as starting material for engineering of an enzyme that may prove useful to industry and should assist in the fundamental understanding of the mechanism by which glycosyl hydrolases evolve product tolerance. | en_US |
| dc.identifier.apacitation | Ndata, K., Nevondo, W., Cekuse, B., van Zyl, Leonardo J., & Trindade, M. (2021). Characterization of a highly xylose tolerant β-xylosidase isolated from high temperature horse manure compost. <i>BMC Biotechnology</i>, 21(Article number: 61), http://hdl.handle.net/11427/35306 | en_ZA |
| dc.identifier.chicagocitation | Ndata, Kanyisa, Walter Nevondo, Bongi Cekuse, Leonardo J. van Zyl, and Marla Trindade "Characterization of a highly xylose tolerant β-xylosidase isolated from high temperature horse manure compost." <i>BMC Biotechnology</i> 21, Article number: 61. (2021) http://hdl.handle.net/11427/35306 | en_ZA |
| dc.identifier.citation | Ndata, K., Nevondo, W., Cekuse, B., van Zyl, Leonardo J. & Trindade, M. 2021. Characterization of a highly xylose tolerant β-xylosidase isolated from high temperature horse manure compost. <i>BMC Biotechnology.</i> 21(Article number: 61) http://hdl.handle.net/11427/35306 | en_ZA |
| dc.identifier.ris | TY - Journal Article AU - Ndata, Kanyisa AU - Nevondo, Walter AU - Cekuse, Bongi AU - van Zyl, Leonardo J. AU - Trindade, Marla AB - Background There is a continued need for improved enzymes for industry. β-xylosidases are enzymes employed in a variety of industries and although many wild-type and engineered variants have been described, enzymes that are highly tolerant of the products produced by catalysis are not readily available and the fundamental mechanisms of tolerance are not well understood. Results Screening of a metagenomic library constructed of mDNA isolated from horse manure compost for β-xylosidase activity identified 26 positive hits. The fosmid clones were sequenced and bioinformatic analysis performed to identity putative β-xylosidases. Based on the novelty of its amino acid sequence and potential thermostability one enzyme (XylP81) was selected for expression and further characterization. XylP81 belongs to the family 39 β-xylosidases, a comparatively rarely found and characterized GH family. The enzyme displayed biochemical characteristics (KM—5.3 mM; Vmax—122 U/mg; kcat—107; Topt—50 °C; pHopt—6) comparable to previously characterized glycoside hydrolase family 39 (GH39) β-xylosidases and despite nucleotide identity to thermophilic species, the enzyme displayed only moderate thermostability with a half-life of 32 min at 60 °C. Apart from acting on substrates predicted for β-xylosidase (xylobiose and 4-nitrophenyl-β-D-xylopyranoside) the enzyme also displayed measurable α-L-arabainofuranosidase, β-galactosidase and β-glucosidase activity. A remarkable feature of this enzyme is its ability to tolerate high concentrations of xylose with a Ki of 1.33 M, a feature that is highly desirable for commercial applications. Conclusions Here we describe a novel β-xylosidase from a poorly studied glycosyl hydrolase family (GH39) which despite having overall kinetic properties similar to other bacterial GH39 β-xylosidases, displays unusually high product tolerance. This trait is shared with only one other member of the GH39 family, the recently described β-xylosidases from Dictyoglomus thermophilum. This feature should allow its use as starting material for engineering of an enzyme that may prove useful to industry and should assist in the fundamental understanding of the mechanism by which glycosyl hydrolases evolve product tolerance. DA - 2021-10-24 DB - OpenUCT DP - University of Cape Town IS - Article number: 61 J1 - BMC Biotechnology KW - Metagenomics KW - Glycoside hydrolase 39 KW - Lignocellulose KW - β-xylosidase LK - https://open.uct.ac.za PY - 2021 T1 - Characterization of a highly xylose tolerant β-xylosidase isolated from high temperature horse manure compost TI - Characterization of a highly xylose tolerant β-xylosidase isolated from high temperature horse manure compost UR - http://hdl.handle.net/11427/35306 ER - | en_ZA |
| dc.identifier.uri | https://doi.org/10.1186/s12896-021-00722-6 | |
| dc.identifier.uri | http://hdl.handle.net/11427/35306 | |
| dc.identifier.vancouvercitation | Ndata K, Nevondo W, Cekuse B, van Zyl Leonardo J, Trindade M. Characterization of a highly xylose tolerant β-xylosidase isolated from high temperature horse manure compost. BMC Biotechnology. 2021;21(Article number: 61) http://hdl.handle.net/11427/35306. | en_ZA |
| dc.language.iso | en | en_US |
| dc.language.rfc3066 | en | |
| dc.publisher.department | Institute of Infectious Disease and Molecular Medicine | en_US |
| dc.publisher.faculty | Faculty of Health Sciences | en_US |
| dc.rights.holder | The Author(s) | |
| dc.rights.uri | http://creativecommons.org/licenses/by/4.0/ | en_US |
| dc.source | BMC Biotechnology | en_US |
| dc.source.journalissue | Article number: 61 | en_US |
| dc.source.journalvolume | 21 | en_US |
| dc.source.uri | https://bmcbiotechnol.biomedcentral.com/ | |
| dc.subject | Metagenomics | en_US |
| dc.subject | Glycoside hydrolase 39 | en_US |
| dc.subject | Lignocellulose | en_US |
| dc.subject | β-xylosidase | en_US |
| dc.title | Characterization of a highly xylose tolerant β-xylosidase isolated from high temperature horse manure compost | en_US |
| dc.type | Journal Article | en_US |