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Production of a high-efficiency cellulase complex via β-glucosidase engineering in Penicillium oxalicum

BACKGROUND: Trichoderma reesei is a widely used model cellulolytic fungus, supplying a highly effective cellulase production system. Recently, the biofuel industry discovered filamentous fungi from the Penicillium genus as a promising alternative to T. reesei. RESULTS: In our study, we present a sys...

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Autores principales: Yao, Guangshan, Wu, Ruimei, Kan, Qinbiao, Gao, Liwei, Liu, Meng, Yang, Piao, Du, Jian, Li, Zhonghai, Qu, Yinbo
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4815182/
https://www.ncbi.nlm.nih.gov/pubmed/27034716
http://dx.doi.org/10.1186/s13068-016-0491-4
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author Yao, Guangshan
Wu, Ruimei
Kan, Qinbiao
Gao, Liwei
Liu, Meng
Yang, Piao
Du, Jian
Li, Zhonghai
Qu, Yinbo
author_facet Yao, Guangshan
Wu, Ruimei
Kan, Qinbiao
Gao, Liwei
Liu, Meng
Yang, Piao
Du, Jian
Li, Zhonghai
Qu, Yinbo
author_sort Yao, Guangshan
collection PubMed
description BACKGROUND: Trichoderma reesei is a widely used model cellulolytic fungus, supplying a highly effective cellulase production system. Recently, the biofuel industry discovered filamentous fungi from the Penicillium genus as a promising alternative to T. reesei. RESULTS: In our study, we present a systematic over-expression analysis of nine β-glucosidase encoding genes in the wild-type strain 114-2 of Penicillium oxalicum. We found that the over-expression of BGL1, BGL4, or BGL5 significantly enhanced both β-glucosidase activity and hydrolysis efficiency of the enzyme system on filter paper. We utilised two strategies to over-express β-glucosidase in the strain RE-10 that—although over-producing cellulase, does so at the cost of the cellulase mixture deficiency. The constitutive promoter of gene pde_02864 encoding 40S ribosomal protein S8 was used to over-express three β-glucosidases: BGL1, BGL4, and BGL5. We found that all mutants show significantly enhanced levels of β-glucosidase at transcriptional, protein, and activity levels. Furthermore, the inducible promoter from bgl2 was used to conditionally over-express the β-glucosidases BGL1 and BGL4. Surprisingly, this induced expression strategy enables significantly improved expression efficiency. The BGL1 over-expressing mutant I1-13 particularly improved the β-glucosidase activity at a factor of 65-folds, resulting in levels of up to 150 U/ml. All our BGL over-expression mutants displayed significant enhancement of cellulolytic ability on both microcrystalline cellulose and filter paper. In addition, they substantially reduced the enzyme loads in the saccharification of a natural lignocellulose material delignified corncob residue (DCCR). The mutant I4-32 with over-expression of BGL4 achieved the highest glucose yield in the saccharification of DCCR at only 25 % enzyme load compared to the parental strain RE-10. CONCLUSIONS: In summary, genetically engineering P. oxalicum to significantly improve β-glucosidase activity is a potent strategy to substantially boost the hydrolytic efficiency of the cellulase cocktail, which will ultimately lead to a considerable reduction of cost for biomass-based biofuel. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13068-016-0491-4) contains supplementary material, which is available to authorized users.
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spelling pubmed-48151822016-04-01 Production of a high-efficiency cellulase complex via β-glucosidase engineering in Penicillium oxalicum Yao, Guangshan Wu, Ruimei Kan, Qinbiao Gao, Liwei Liu, Meng Yang, Piao Du, Jian Li, Zhonghai Qu, Yinbo Biotechnol Biofuels Research BACKGROUND: Trichoderma reesei is a widely used model cellulolytic fungus, supplying a highly effective cellulase production system. Recently, the biofuel industry discovered filamentous fungi from the Penicillium genus as a promising alternative to T. reesei. RESULTS: In our study, we present a systematic over-expression analysis of nine β-glucosidase encoding genes in the wild-type strain 114-2 of Penicillium oxalicum. We found that the over-expression of BGL1, BGL4, or BGL5 significantly enhanced both β-glucosidase activity and hydrolysis efficiency of the enzyme system on filter paper. We utilised two strategies to over-express β-glucosidase in the strain RE-10 that—although over-producing cellulase, does so at the cost of the cellulase mixture deficiency. The constitutive promoter of gene pde_02864 encoding 40S ribosomal protein S8 was used to over-express three β-glucosidases: BGL1, BGL4, and BGL5. We found that all mutants show significantly enhanced levels of β-glucosidase at transcriptional, protein, and activity levels. Furthermore, the inducible promoter from bgl2 was used to conditionally over-express the β-glucosidases BGL1 and BGL4. Surprisingly, this induced expression strategy enables significantly improved expression efficiency. The BGL1 over-expressing mutant I1-13 particularly improved the β-glucosidase activity at a factor of 65-folds, resulting in levels of up to 150 U/ml. All our BGL over-expression mutants displayed significant enhancement of cellulolytic ability on both microcrystalline cellulose and filter paper. In addition, they substantially reduced the enzyme loads in the saccharification of a natural lignocellulose material delignified corncob residue (DCCR). The mutant I4-32 with over-expression of BGL4 achieved the highest glucose yield in the saccharification of DCCR at only 25 % enzyme load compared to the parental strain RE-10. CONCLUSIONS: In summary, genetically engineering P. oxalicum to significantly improve β-glucosidase activity is a potent strategy to substantially boost the hydrolytic efficiency of the cellulase cocktail, which will ultimately lead to a considerable reduction of cost for biomass-based biofuel. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13068-016-0491-4) contains supplementary material, which is available to authorized users. BioMed Central 2016-03-31 /pmc/articles/PMC4815182/ /pubmed/27034716 http://dx.doi.org/10.1186/s13068-016-0491-4 Text en © Yao et al. 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
Yao, Guangshan
Wu, Ruimei
Kan, Qinbiao
Gao, Liwei
Liu, Meng
Yang, Piao
Du, Jian
Li, Zhonghai
Qu, Yinbo
Production of a high-efficiency cellulase complex via β-glucosidase engineering in Penicillium oxalicum
title Production of a high-efficiency cellulase complex via β-glucosidase engineering in Penicillium oxalicum
title_full Production of a high-efficiency cellulase complex via β-glucosidase engineering in Penicillium oxalicum
title_fullStr Production of a high-efficiency cellulase complex via β-glucosidase engineering in Penicillium oxalicum
title_full_unstemmed Production of a high-efficiency cellulase complex via β-glucosidase engineering in Penicillium oxalicum
title_short Production of a high-efficiency cellulase complex via β-glucosidase engineering in Penicillium oxalicum
title_sort production of a high-efficiency cellulase complex via β-glucosidase engineering in penicillium oxalicum
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4815182/
https://www.ncbi.nlm.nih.gov/pubmed/27034716
http://dx.doi.org/10.1186/s13068-016-0491-4
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