Presence of protein production enhancers results in significantly higher methanol-induced protein production in Pichia pastoris
Abstract Background The yeast Komagataella phaffii, better known as Pichia pastoris, is a commonly used host for recombinant protein production. Here expression vectors are reported that address the different steps of the transcription–translation–secretion pathway of heterologous protein production...
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| Format: | Article |
| Language: | English |
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BMC
2018-07-01
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| Series: | Microbial Cell Factories |
| Subjects: | |
| Online Access: | http://link.springer.com/article/10.1186/s12934-018-0961-4 |
| _version_ | 1818450553259163648 |
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| author | Loknath Gidijala Stefan Uthoff Sebastiaan J. van Kampen Alexander Steinbüchel Raymond M. D. Verhaert |
| author_facet | Loknath Gidijala Stefan Uthoff Sebastiaan J. van Kampen Alexander Steinbüchel Raymond M. D. Verhaert |
| author_sort | Loknath Gidijala |
| collection | DOAJ |
| description | Abstract Background The yeast Komagataella phaffii, better known as Pichia pastoris, is a commonly used host for recombinant protein production. Here expression vectors are reported that address the different steps of the transcription–translation–secretion pathway of heterologous protein production. Results Transcription and translation enhancing elements were introduced in an expression cassette for the production of recombinant Aspergillus niger feruloyl esterase A. The yield was increased by threefold as compared to the yield without these elements. Multiple copy strains were selected using a zeocin resistance marker in the expression cassette and showed another sixfold higher yield. Modification of the C-terminal amino acid sequence of the secretion signal did not significantly improve the production yield. Similar data were obtained for the production of another protein, recombinant human interleukin 8. Upscaling to fed-batch fermentation conditions resulted in a twofold increase for reference strains, while for strains with enhancing elements a tenfold improvement was observed. Conclusions Pichia pastoris is used for recombinant protein production in industrial fermentations. By addressing the transcription and translation of mRNA coding for recombinant protein, significant yield improvement was obtained. The yield improvement obtained under microscale conditions was maintained under fed-batch fermentation conditions. These data demonstrate the potential of these expression vectors for large scale application as improved production of proteins has major implications on the economics and sustainability of biocatalyst dependent production processes e.g. for the production of pharmaceuticals and for the bioconversions of complex molecules. |
| first_indexed | 2024-12-14T20:53:08Z |
| format | Article |
| id | doaj.art-8ec521d7dd704f9ba80f59a964c21571 |
| institution | Directory Open Access Journal |
| issn | 1475-2859 |
| language | English |
| last_indexed | 2024-12-14T20:53:08Z |
| publishDate | 2018-07-01 |
| publisher | BMC |
| record_format | Article |
| series | Microbial Cell Factories |
| spelling | doaj.art-8ec521d7dd704f9ba80f59a964c215712022-12-21T22:47:45ZengBMCMicrobial Cell Factories1475-28592018-07-0117111110.1186/s12934-018-0961-4Presence of protein production enhancers results in significantly higher methanol-induced protein production in Pichia pastorisLoknath Gidijala0Stefan Uthoff1Sebastiaan J. van Kampen2Alexander Steinbüchel3Raymond M. D. Verhaert4ProteoNic BVInstitut für Molekulare Mikrobiologie und Biotechnologie, Westfälische Wilhelms-Universität MünsterProteoNic BVInstitut für Molekulare Mikrobiologie und Biotechnologie, Westfälische Wilhelms-Universität MünsterProteoNic BVAbstract Background The yeast Komagataella phaffii, better known as Pichia pastoris, is a commonly used host for recombinant protein production. Here expression vectors are reported that address the different steps of the transcription–translation–secretion pathway of heterologous protein production. Results Transcription and translation enhancing elements were introduced in an expression cassette for the production of recombinant Aspergillus niger feruloyl esterase A. The yield was increased by threefold as compared to the yield without these elements. Multiple copy strains were selected using a zeocin resistance marker in the expression cassette and showed another sixfold higher yield. Modification of the C-terminal amino acid sequence of the secretion signal did not significantly improve the production yield. Similar data were obtained for the production of another protein, recombinant human interleukin 8. Upscaling to fed-batch fermentation conditions resulted in a twofold increase for reference strains, while for strains with enhancing elements a tenfold improvement was observed. Conclusions Pichia pastoris is used for recombinant protein production in industrial fermentations. By addressing the transcription and translation of mRNA coding for recombinant protein, significant yield improvement was obtained. The yield improvement obtained under microscale conditions was maintained under fed-batch fermentation conditions. These data demonstrate the potential of these expression vectors for large scale application as improved production of proteins has major implications on the economics and sustainability of biocatalyst dependent production processes e.g. for the production of pharmaceuticals and for the bioconversions of complex molecules.http://link.springer.com/article/10.1186/s12934-018-0961-4Recombinant protein productionYield enhancementPichia pastorisYeastGene expression |
| spellingShingle | Loknath Gidijala Stefan Uthoff Sebastiaan J. van Kampen Alexander Steinbüchel Raymond M. D. Verhaert Presence of protein production enhancers results in significantly higher methanol-induced protein production in Pichia pastoris Microbial Cell Factories Recombinant protein production Yield enhancement Pichia pastoris Yeast Gene expression |
| title | Presence of protein production enhancers results in significantly higher methanol-induced protein production in Pichia pastoris |
| title_full | Presence of protein production enhancers results in significantly higher methanol-induced protein production in Pichia pastoris |
| title_fullStr | Presence of protein production enhancers results in significantly higher methanol-induced protein production in Pichia pastoris |
| title_full_unstemmed | Presence of protein production enhancers results in significantly higher methanol-induced protein production in Pichia pastoris |
| title_short | Presence of protein production enhancers results in significantly higher methanol-induced protein production in Pichia pastoris |
| title_sort | presence of protein production enhancers results in significantly higher methanol induced protein production in pichia pastoris |
| topic | Recombinant protein production Yield enhancement Pichia pastoris Yeast Gene expression |
| url | http://link.springer.com/article/10.1186/s12934-018-0961-4 |
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