Expressing accessory proteins in cellulolytic Yarrowia lipolytica to improve the conversion yield of recalcitrant cellulose
| Autor: | Zhongpeng Guo, Michael J. O’Donohue, Alain Marty, Sophie Bozonnet, Sophie Duquesne, Jean-Marc Nicaud |
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| Přispěvatelé: | Laboratoire d'Ingénierie des Systèmes Biologiques et des Procédés (LISBP), Centre National de la Recherche Scientifique (CNRS)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National de la Recherche Agronomique (INRA), MICrobiologie de l'ALImentation au Service de la Santé (MICALIS), Institut National de la Recherche Agronomique (INRA)-AgroParisTech, Agence Nationale de la Recherche (Investissements d'Avenir program) [ANR-11-BTBR-0003], Institut National de la Recherche Agronomique (INRA)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS), Guo, Zhongpeng, Marty, Alain, O'Donohue, Michael |
| Jazyk: | angličtina |
| Rok vydání: | 2017 |
| Předmět: |
0106 biological sciences
0301 basic medicine enzyme cellulolytique [SDV.BIO]Life Sciences [q-bio]/Biotechnology hydrolyse enzymatique 01 natural sciences Applied Microbiology and Biotechnology xylan xylane chemistry.chemical_compound Trichoderma reesei Swollenin 2. Zero hunger chemistry.chemical_classification biology activité hydrolytique glucan cellulose General Energy Biochemistry Xylanase Biotechnology glucane lcsh:Biotechnology Cellulase Biotechnologies Management Monitoring Policy and Law Polysaccharide lcsh:Fuel 03 medical and health sciences Consolidated bioprocessing Accessory proteins lcsh:TP315-360 010608 biotechnology Enzymatic hydrolysis lcsh:TP248.13-248.65 Cellulolytic biocatalyst Cellulose Lytic polysaccharide monooxygenase Renewable Energy Sustainability and the Environment Research enzymatic hydrolysis Yarrowia biology.organism_classification Xylan 030104 developmental biology chemistry Yarrowia lipolytica cellulolytic biocatalyst consolidated bioprocessing accessory proteins xylanase lytic polysaccharide monooxygenase swollenin biology.protein yarrowia lipolytica |
| Zdroj: | Biotechnology for Biofuels Biotechnology for Biofuels, BioMed Central, 2017, 10 (1), 16 p. ⟨10.1186/s13068-017-0990-y⟩ Biotechnology for Biofuels, Vol 10, Iss 1, Pp 1-16 (2017) Biotechnology for Biofuels, 2017, 10 (1), 16 p. ⟨10.1186/s13068-017-0990-y⟩ Biotechnology for Biofuels 1 (10), 16 p.. (2017) |
| ISSN: | 1754-6834 |
| DOI: | 10.1186/s13068-017-0990-y⟩ |
| Popis: | Background A recently constructed cellulolytic Yarrowia lipolytica is able to grow efficiently on an industrial organosolv cellulose pulp, but shows limited ability to degrade crystalline cellulose. In this work, we have further engineered this strain, adding accessory proteins xylanase II (XYNII), lytic polysaccharide monooxygenase (LPMO), and swollenin (SWO) from Trichoderma reesei in order to enhance the degradation of recalcitrant substrate. Results The production of EG I was enhanced using a promoter engineering strategy. This provided a new cellulolytic Y. lipolytica strain, which compared to the parent strain, exhibited higher hydrolytic activity on different cellulosic substrates. Furthermore, three accessory proteins, TrXYNII, TrLPMOA and TrSWO, were individually expressed in cellulolytic and non-cellulolytic Y. lipolytica. The amount of rhTrXYNII and rhTrLPMOA secreted by non-cellulolytic Y. lipolytica in YTD medium during batch cultivation in flasks was approximately 62 and 52 mg/L, respectively. The purified rhTrXYNII showed a specific activity of 532 U/mg-protein on beechwood xylan, while rhTrLPMOA exhibited a specific activity of 14.4 U/g-protein when using the Amplex Red/horseradish peroxidase assay. Characterization of rhTrLPMOA revealed that this protein displays broad specificity against β-(1,4)-linked glucans, but is inactive on xylan. Further studies showed that the presence of TrLPMOA synergistically enhanced enzymatic hydrolysis of cellulose by cellulases, while TrSWO1 boosted cellulose hydrolysis only when it was applied before the action of cellulases. The presence of rTrXYNII enhanced enzymatic hydrolysis of an industrial cellulose pulp and of wheat straw. Co-expressing TrXYNII and TrLPMOA in cellulolytic Y. lipolytica with enhanced EG I production procured a novel engineered Y. lipolytica strain that displayed enhanced ability to degrade both amorphous (CIMV-cellulose) and recalcitrant crystalline cellulose in complex biomass (wheat straw) by 16 and 90%, respectively. Conclusions This study has provided a potent cellulose-degrading Y. lipolytica strain that co-expresses a core set of cellulolytic enzymes and some accessory proteins. Results reveal that the tuning of cellulase production and the production of accessory proteins leads to optimized performance. Accordingly, the beneficial effect of accessory proteins for cellulase-mediated degradation of cellulose is underlined, especially when crystalline cellulose and complex biomass are used as substrates. Findings specifically underline the benefits and specific properties of swollenin. Although in our study swollenin clearly promoted cellulase action, its use requires process redesign to accommodate its specific mode of action. Electronic supplementary material The online version of this article (10.1186/s13068-017-0990-y) contains supplementary material, which is available to authorized users. |
| Databáze: | OpenAIRE |
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