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The GH51 α-l-arabinofuranosidase from Paenibacillus sp. THS1 is multifunctional, hydrolyzing main-chain and side-chain glycosidic bonds in heteroxylans

BACKGROUND: Conceptually, multi-functional enzymes are attractive because in the case of complex polymer hydrolysis having two or more activities defined by a single enzyme offers the possibility of synergy and reduced enzyme cocktail complexity. Nevertheless, multi-functional enzymes are quite rare...

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Autores principales: Bouraoui, Hanen, Desrousseaux, Marie-Laure, Ioannou, Eleni, Alvira, Pablo, Manaï, Mohamed, Rémond, Caroline, Dumon, Claire, Fernandez-Fuentes, Narcis, O’Donohue, Michael J.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4939007/
https://www.ncbi.nlm.nih.gov/pubmed/27398094
http://dx.doi.org/10.1186/s13068-016-0550-x
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author Bouraoui, Hanen
Desrousseaux, Marie-Laure
Ioannou, Eleni
Alvira, Pablo
Manaï, Mohamed
Rémond, Caroline
Dumon, Claire
Fernandez-Fuentes, Narcis
O’Donohue, Michael J.
author_facet Bouraoui, Hanen
Desrousseaux, Marie-Laure
Ioannou, Eleni
Alvira, Pablo
Manaï, Mohamed
Rémond, Caroline
Dumon, Claire
Fernandez-Fuentes, Narcis
O’Donohue, Michael J.
author_sort Bouraoui, Hanen
collection PubMed
description BACKGROUND: Conceptually, multi-functional enzymes are attractive because in the case of complex polymer hydrolysis having two or more activities defined by a single enzyme offers the possibility of synergy and reduced enzyme cocktail complexity. Nevertheless, multi-functional enzymes are quite rare and are generally multi-domain assemblies with each activity being defined by a separate protein module. However, a recent report described a GH51 arabinofuranosidase from Alicyclobacillus sp. A4 that displays both α-l-arabinofuranosidase and β-d-xylanase activities, which are defined by a single active site. Following on from this, we describe in detail another multi-functional GH51 arabinofuranosidase and discuss the molecular basis of multifunctionality. RESULTS: THSAbf is a GH51 α-l-arabinofuranosidase. Characterization revealed that THSAbf is active up to 75 °C, stable at 60 °C and active over a broad pH range (4–7). THSAbf preferentially releases para-nitrophenyl from the l-arabinofuranoside (k(cat)/K(M) = 1050 s(−1) mM(−1)) and to some extent from d-galactofuranoside and d-xyloside. THSAbf is active on 4-O-methylglucuronoxylans from birch and beechwood (10.8 and 14.4 U mg(−1), respectively) and on sugar beet branched and linear arabinans (1.1 ± 0.24 and 1.8 ± 0.1 U mg(−1)). Further investigation revealed that like the Alicyclobacillus sp. A4 α-l-arabinofuranosidase, THSAbf also displays endo-xylanase activity, cleaving β-1,4 bonds in heteroxylans. The optimum pH for THASAbf activity is substrate dependent, but ablation of the catalytic nucleophile caused a general loss of activity, indicating the involvement of a single active center. Combining the α-l-arabinofuranosidase with a GH11 endoxylanase did not procure synergy. The molecular modeling of THSAbf revealed a wide active site cleft and clues to explain multi-functionality. CONCLUSION: The discovery of single active site, multifunctional enzymes such as THSAbf opens up exciting avenues for enzyme engineering and the development of new biomass-degrading cocktails that could considerably reduce enzyme production costs. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13068-016-0550-x) contains supplementary material, which is available to authorized users.
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spelling pubmed-49390072016-07-10 The GH51 α-l-arabinofuranosidase from Paenibacillus sp. THS1 is multifunctional, hydrolyzing main-chain and side-chain glycosidic bonds in heteroxylans Bouraoui, Hanen Desrousseaux, Marie-Laure Ioannou, Eleni Alvira, Pablo Manaï, Mohamed Rémond, Caroline Dumon, Claire Fernandez-Fuentes, Narcis O’Donohue, Michael J. Biotechnol Biofuels Research BACKGROUND: Conceptually, multi-functional enzymes are attractive because in the case of complex polymer hydrolysis having two or more activities defined by a single enzyme offers the possibility of synergy and reduced enzyme cocktail complexity. Nevertheless, multi-functional enzymes are quite rare and are generally multi-domain assemblies with each activity being defined by a separate protein module. However, a recent report described a GH51 arabinofuranosidase from Alicyclobacillus sp. A4 that displays both α-l-arabinofuranosidase and β-d-xylanase activities, which are defined by a single active site. Following on from this, we describe in detail another multi-functional GH51 arabinofuranosidase and discuss the molecular basis of multifunctionality. RESULTS: THSAbf is a GH51 α-l-arabinofuranosidase. Characterization revealed that THSAbf is active up to 75 °C, stable at 60 °C and active over a broad pH range (4–7). THSAbf preferentially releases para-nitrophenyl from the l-arabinofuranoside (k(cat)/K(M) = 1050 s(−1) mM(−1)) and to some extent from d-galactofuranoside and d-xyloside. THSAbf is active on 4-O-methylglucuronoxylans from birch and beechwood (10.8 and 14.4 U mg(−1), respectively) and on sugar beet branched and linear arabinans (1.1 ± 0.24 and 1.8 ± 0.1 U mg(−1)). Further investigation revealed that like the Alicyclobacillus sp. A4 α-l-arabinofuranosidase, THSAbf also displays endo-xylanase activity, cleaving β-1,4 bonds in heteroxylans. The optimum pH for THASAbf activity is substrate dependent, but ablation of the catalytic nucleophile caused a general loss of activity, indicating the involvement of a single active center. Combining the α-l-arabinofuranosidase with a GH11 endoxylanase did not procure synergy. The molecular modeling of THSAbf revealed a wide active site cleft and clues to explain multi-functionality. CONCLUSION: The discovery of single active site, multifunctional enzymes such as THSAbf opens up exciting avenues for enzyme engineering and the development of new biomass-degrading cocktails that could considerably reduce enzyme production costs. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13068-016-0550-x) contains supplementary material, which is available to authorized users. BioMed Central 2016-07-08 /pmc/articles/PMC4939007/ /pubmed/27398094 http://dx.doi.org/10.1186/s13068-016-0550-x Text en © The Author(s) 2016 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
spellingShingle Research
Bouraoui, Hanen
Desrousseaux, Marie-Laure
Ioannou, Eleni
Alvira, Pablo
Manaï, Mohamed
Rémond, Caroline
Dumon, Claire
Fernandez-Fuentes, Narcis
O’Donohue, Michael J.
The GH51 α-l-arabinofuranosidase from Paenibacillus sp. THS1 is multifunctional, hydrolyzing main-chain and side-chain glycosidic bonds in heteroxylans
title The GH51 α-l-arabinofuranosidase from Paenibacillus sp. THS1 is multifunctional, hydrolyzing main-chain and side-chain glycosidic bonds in heteroxylans
title_full The GH51 α-l-arabinofuranosidase from Paenibacillus sp. THS1 is multifunctional, hydrolyzing main-chain and side-chain glycosidic bonds in heteroxylans
title_fullStr The GH51 α-l-arabinofuranosidase from Paenibacillus sp. THS1 is multifunctional, hydrolyzing main-chain and side-chain glycosidic bonds in heteroxylans
title_full_unstemmed The GH51 α-l-arabinofuranosidase from Paenibacillus sp. THS1 is multifunctional, hydrolyzing main-chain and side-chain glycosidic bonds in heteroxylans
title_short The GH51 α-l-arabinofuranosidase from Paenibacillus sp. THS1 is multifunctional, hydrolyzing main-chain and side-chain glycosidic bonds in heteroxylans
title_sort gh51 α-l-arabinofuranosidase from paenibacillus sp. ths1 is multifunctional, hydrolyzing main-chain and side-chain glycosidic bonds in heteroxylans
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4939007/
https://www.ncbi.nlm.nih.gov/pubmed/27398094
http://dx.doi.org/10.1186/s13068-016-0550-x
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