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Conversion of xylan by recyclable spores of Bacillus subtilis displaying thermophilic enzymes

BACKGROUND: The Bacillus subtilis spore has long been used to display antigens and enzymes. Spore display can be accomplished by a recombinant and a non-recombinant approach, with the latter proved more efficient than the recombinant one. We used the non-recombinant approach to independently adsorb...

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Autores principales: Mattossovich, Rosanna, Iacono, Roberta, Cangiano, Giuseppina, Cobucci-Ponzano, Beatrice, Isticato, Rachele, Moracci, Marco, Ricca, Ezio
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5706412/
https://www.ncbi.nlm.nih.gov/pubmed/29183330
http://dx.doi.org/10.1186/s12934-017-0833-3
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author Mattossovich, Rosanna
Iacono, Roberta
Cangiano, Giuseppina
Cobucci-Ponzano, Beatrice
Isticato, Rachele
Moracci, Marco
Ricca, Ezio
author_facet Mattossovich, Rosanna
Iacono, Roberta
Cangiano, Giuseppina
Cobucci-Ponzano, Beatrice
Isticato, Rachele
Moracci, Marco
Ricca, Ezio
author_sort Mattossovich, Rosanna
collection PubMed
description BACKGROUND: The Bacillus subtilis spore has long been used to display antigens and enzymes. Spore display can be accomplished by a recombinant and a non-recombinant approach, with the latter proved more efficient than the recombinant one. We used the non-recombinant approach to independently adsorb two thermophilic enzymes, GH10-XA, an endo-1,4-β-xylanase (EC 3.2.1.8) from Alicyclobacillus acidocaldarius, and GH3-XT, a β-xylosidase (EC 3.2.1.37) from Thermotoga thermarum. These enzymes catalyze, respectively, the endohydrolysis of (1-4)-β-d-xylosidic linkages of xylans and the hydrolysis of (1-4)-β-d-xylans to remove successive d-xylose residues from the non-reducing termini. RESULTS: We report that both purified enzymes were independently adsorbed on purified spores of B. subtilis. The adsorption was tight and both enzymes retained part of their specific activity. When spores displaying either GH10-XA or GH3-XT were mixed together, xylan was hydrolysed more efficiently than by a mixture of the two free, not spore-adsorbed, enzymes. The high total activity of the spore-bound enzymes is most likely due to a stabilization of the enzymes that, upon adsorption on the spore, remained active at the reaction conditions for longer than the free enzymes. Spore-adsorbed enzymes, collected after the two-step reaction and incubated with fresh substrate, were still active and able to continue xylan degradation. The recycling of the mixed spore-bound enzymes allowed a strong increase of xylan degradation. CONCLUSION: Our results indicate that the two-step degradation of xylans can be accomplished by mixing spores displaying either one of two required enzymes. The two-step process occurs more efficiently than with the two un-adsorbed, free enzymes and adsorbed spores can be reused for at least one other reaction round. The efficiency of the process, the reusability of the adsorbed enzymes, and the well documented robustness of spores of B. subtilis indicate the spore as a suitable platform to display enzymes for single as well as multi-step reactions.
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spelling pubmed-57064122017-12-06 Conversion of xylan by recyclable spores of Bacillus subtilis displaying thermophilic enzymes Mattossovich, Rosanna Iacono, Roberta Cangiano, Giuseppina Cobucci-Ponzano, Beatrice Isticato, Rachele Moracci, Marco Ricca, Ezio Microb Cell Fact Research BACKGROUND: The Bacillus subtilis spore has long been used to display antigens and enzymes. Spore display can be accomplished by a recombinant and a non-recombinant approach, with the latter proved more efficient than the recombinant one. We used the non-recombinant approach to independently adsorb two thermophilic enzymes, GH10-XA, an endo-1,4-β-xylanase (EC 3.2.1.8) from Alicyclobacillus acidocaldarius, and GH3-XT, a β-xylosidase (EC 3.2.1.37) from Thermotoga thermarum. These enzymes catalyze, respectively, the endohydrolysis of (1-4)-β-d-xylosidic linkages of xylans and the hydrolysis of (1-4)-β-d-xylans to remove successive d-xylose residues from the non-reducing termini. RESULTS: We report that both purified enzymes were independently adsorbed on purified spores of B. subtilis. The adsorption was tight and both enzymes retained part of their specific activity. When spores displaying either GH10-XA or GH3-XT were mixed together, xylan was hydrolysed more efficiently than by a mixture of the two free, not spore-adsorbed, enzymes. The high total activity of the spore-bound enzymes is most likely due to a stabilization of the enzymes that, upon adsorption on the spore, remained active at the reaction conditions for longer than the free enzymes. Spore-adsorbed enzymes, collected after the two-step reaction and incubated with fresh substrate, were still active and able to continue xylan degradation. The recycling of the mixed spore-bound enzymes allowed a strong increase of xylan degradation. CONCLUSION: Our results indicate that the two-step degradation of xylans can be accomplished by mixing spores displaying either one of two required enzymes. The two-step process occurs more efficiently than with the two un-adsorbed, free enzymes and adsorbed spores can be reused for at least one other reaction round. The efficiency of the process, the reusability of the adsorbed enzymes, and the well documented robustness of spores of B. subtilis indicate the spore as a suitable platform to display enzymes for single as well as multi-step reactions. BioMed Central 2017-11-28 /pmc/articles/PMC5706412/ /pubmed/29183330 http://dx.doi.org/10.1186/s12934-017-0833-3 Text en © The Author(s) 2017 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
Mattossovich, Rosanna
Iacono, Roberta
Cangiano, Giuseppina
Cobucci-Ponzano, Beatrice
Isticato, Rachele
Moracci, Marco
Ricca, Ezio
Conversion of xylan by recyclable spores of Bacillus subtilis displaying thermophilic enzymes
title Conversion of xylan by recyclable spores of Bacillus subtilis displaying thermophilic enzymes
title_full Conversion of xylan by recyclable spores of Bacillus subtilis displaying thermophilic enzymes
title_fullStr Conversion of xylan by recyclable spores of Bacillus subtilis displaying thermophilic enzymes
title_full_unstemmed Conversion of xylan by recyclable spores of Bacillus subtilis displaying thermophilic enzymes
title_short Conversion of xylan by recyclable spores of Bacillus subtilis displaying thermophilic enzymes
title_sort conversion of xylan by recyclable spores of bacillus subtilis displaying thermophilic enzymes
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5706412/
https://www.ncbi.nlm.nih.gov/pubmed/29183330
http://dx.doi.org/10.1186/s12934-017-0833-3
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