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Displaying Lipase B from Candida antarctica in Pichia pastoris Using the Yeast Surface Display Approach: Prospection of a New Anchor and Characterization of the Whole Cell Biocatalyst
Yeast Surface Display (YSD) is a strategy to anchor proteins on the yeast cell wall which has been employed to increase enzyme stability thus decreasing production costs. Lipase B from Candida antarctica (LipB) is one of the most studied enzymes in the context of industrial biotechnology. This study...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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Public Library of Science
2015
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4624902/ https://www.ncbi.nlm.nih.gov/pubmed/26510006 http://dx.doi.org/10.1371/journal.pone.0141454 |
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author | Moura, Marcelo Victor Holanda da Silva, Giulia Pontes Machado, Antônio Carlos de Oliveira Torres, Fernando Araripe Gonçalves Freire, Denise Maria Guimarães Almeida, Rodrigo Volcan |
author_facet | Moura, Marcelo Victor Holanda da Silva, Giulia Pontes Machado, Antônio Carlos de Oliveira Torres, Fernando Araripe Gonçalves Freire, Denise Maria Guimarães Almeida, Rodrigo Volcan |
author_sort | Moura, Marcelo Victor Holanda |
collection | PubMed |
description | Yeast Surface Display (YSD) is a strategy to anchor proteins on the yeast cell wall which has been employed to increase enzyme stability thus decreasing production costs. Lipase B from Candida antarctica (LipB) is one of the most studied enzymes in the context of industrial biotechnology. This study aimed to assess the biochemical features of this important biocatalyst when immobilized on the cell surface of the methylotrophic yeast Pichia pastoris using the YSD approach. For that purpose, two anchors were tested. The first (Flo9) was identified after a prospection of the P. pastoris genome being related to the family of flocculins similar to Flo1 but significantly smaller. The second is the Protein with Internal Repeats (Pir1) from P. pastoris. An immunolocalization assay showed that both anchor proteins were able to display the reporter protein EGFP in the yeast outer cell wall. LipB was expressed in P. pastoris fused either to Flo9 (FLOLIPB) or Pir1 (PIRLIPB). Both constructions showed hydrolytic activity towards tributyrin (>100 U/mg(dcw) and >80 U/mg(dcw,) respectively), optimal hydrolytic activity around 45°C and pH 7.0, higher thermostability at 45°C and stability in organic solvents when compared to a free lipase. |
format | Online Article Text |
id | pubmed-4624902 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2015 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-46249022015-11-06 Displaying Lipase B from Candida antarctica in Pichia pastoris Using the Yeast Surface Display Approach: Prospection of a New Anchor and Characterization of the Whole Cell Biocatalyst Moura, Marcelo Victor Holanda da Silva, Giulia Pontes Machado, Antônio Carlos de Oliveira Torres, Fernando Araripe Gonçalves Freire, Denise Maria Guimarães Almeida, Rodrigo Volcan PLoS One Research Article Yeast Surface Display (YSD) is a strategy to anchor proteins on the yeast cell wall which has been employed to increase enzyme stability thus decreasing production costs. Lipase B from Candida antarctica (LipB) is one of the most studied enzymes in the context of industrial biotechnology. This study aimed to assess the biochemical features of this important biocatalyst when immobilized on the cell surface of the methylotrophic yeast Pichia pastoris using the YSD approach. For that purpose, two anchors were tested. The first (Flo9) was identified after a prospection of the P. pastoris genome being related to the family of flocculins similar to Flo1 but significantly smaller. The second is the Protein with Internal Repeats (Pir1) from P. pastoris. An immunolocalization assay showed that both anchor proteins were able to display the reporter protein EGFP in the yeast outer cell wall. LipB was expressed in P. pastoris fused either to Flo9 (FLOLIPB) or Pir1 (PIRLIPB). Both constructions showed hydrolytic activity towards tributyrin (>100 U/mg(dcw) and >80 U/mg(dcw,) respectively), optimal hydrolytic activity around 45°C and pH 7.0, higher thermostability at 45°C and stability in organic solvents when compared to a free lipase. Public Library of Science 2015-10-28 /pmc/articles/PMC4624902/ /pubmed/26510006 http://dx.doi.org/10.1371/journal.pone.0141454 Text en © 2015 Moura et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited. |
spellingShingle | Research Article Moura, Marcelo Victor Holanda da Silva, Giulia Pontes Machado, Antônio Carlos de Oliveira Torres, Fernando Araripe Gonçalves Freire, Denise Maria Guimarães Almeida, Rodrigo Volcan Displaying Lipase B from Candida antarctica in Pichia pastoris Using the Yeast Surface Display Approach: Prospection of a New Anchor and Characterization of the Whole Cell Biocatalyst |
title | Displaying Lipase B from Candida antarctica in Pichia pastoris Using the Yeast Surface Display Approach: Prospection of a New Anchor and Characterization of the Whole Cell Biocatalyst |
title_full | Displaying Lipase B from Candida antarctica in Pichia pastoris Using the Yeast Surface Display Approach: Prospection of a New Anchor and Characterization of the Whole Cell Biocatalyst |
title_fullStr | Displaying Lipase B from Candida antarctica in Pichia pastoris Using the Yeast Surface Display Approach: Prospection of a New Anchor and Characterization of the Whole Cell Biocatalyst |
title_full_unstemmed | Displaying Lipase B from Candida antarctica in Pichia pastoris Using the Yeast Surface Display Approach: Prospection of a New Anchor and Characterization of the Whole Cell Biocatalyst |
title_short | Displaying Lipase B from Candida antarctica in Pichia pastoris Using the Yeast Surface Display Approach: Prospection of a New Anchor and Characterization of the Whole Cell Biocatalyst |
title_sort | displaying lipase b from candida antarctica in pichia pastoris using the yeast surface display approach: prospection of a new anchor and characterization of the whole cell biocatalyst |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4624902/ https://www.ncbi.nlm.nih.gov/pubmed/26510006 http://dx.doi.org/10.1371/journal.pone.0141454 |
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