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Investigation on the processing and improving the cleavage efficiency of furin cleavage sites in Pichia pastoris

BACKGROUND: Proprotein convertase furin is responsible for the processing of a wide variety of precursors consisted of signal peptide, propeptide and mature peptide in mammal. Many precursors processed by furin have important physiological functions and can be recombinantly expressed in Pichia pasto...

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Autores principales: Huang, Yide, Long, Yanyu, Li, Suhuan, Lin, Ting, Wu, Jingwen, Zhang, Yafei, Lin, Yao
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6223083/
https://www.ncbi.nlm.nih.gov/pubmed/30409181
http://dx.doi.org/10.1186/s12934-018-1020-x
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author Huang, Yide
Long, Yanyu
Li, Suhuan
Lin, Ting
Wu, Jingwen
Zhang, Yafei
Lin, Yao
author_facet Huang, Yide
Long, Yanyu
Li, Suhuan
Lin, Ting
Wu, Jingwen
Zhang, Yafei
Lin, Yao
author_sort Huang, Yide
collection PubMed
description BACKGROUND: Proprotein convertase furin is responsible for the processing of a wide variety of precursors consisted of signal peptide, propeptide and mature peptide in mammal. Many precursors processed by furin have important physiological functions and can be recombinantly expressed in Pichia pastoris expression system for research, pharmaceutical and vaccine applications. However, it is not clear whether the furin cleavage sites between the propeptide and mature peptide can be properly processed in P. pastoris, bringing uncertainty for proper expression of the coding DNA sequences of furin precursors containing the propeptides and mature peptides. RESULTS: In this study, we evaluated the ability of P. pastoris to process furin cleavage sites and how to improve the cleavage efficiencies of furin cleavage sites in P. pastoris. The results showed that P. pastoris can process furin cleavage sites but the cleavage efficiencies are not high. Arg residue at position P1 or P4 in furin cleavage sites significantly affect cleavage efficiency in P. pastoris. Kex2 protease, but not YPS1, in P. pastoris is responsible for processing furin cleavage sites. Heterologous expression of furin or overexpression of Kex2 in P. pastoris effectively increased cleavage efficiencies of furin cleavage sites. CONCLUSIONS: Our investigation on the processing of furin cleavage sites provides important information for recombinant expression of furin precursors in P. pastoris. Furin or Kex2 overexpressing strains may be good choices for expressing precursors processed by furin in P. pastoris.
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spelling pubmed-62230832018-11-19 Investigation on the processing and improving the cleavage efficiency of furin cleavage sites in Pichia pastoris Huang, Yide Long, Yanyu Li, Suhuan Lin, Ting Wu, Jingwen Zhang, Yafei Lin, Yao Microb Cell Fact Research BACKGROUND: Proprotein convertase furin is responsible for the processing of a wide variety of precursors consisted of signal peptide, propeptide and mature peptide in mammal. Many precursors processed by furin have important physiological functions and can be recombinantly expressed in Pichia pastoris expression system for research, pharmaceutical and vaccine applications. However, it is not clear whether the furin cleavage sites between the propeptide and mature peptide can be properly processed in P. pastoris, bringing uncertainty for proper expression of the coding DNA sequences of furin precursors containing the propeptides and mature peptides. RESULTS: In this study, we evaluated the ability of P. pastoris to process furin cleavage sites and how to improve the cleavage efficiencies of furin cleavage sites in P. pastoris. The results showed that P. pastoris can process furin cleavage sites but the cleavage efficiencies are not high. Arg residue at position P1 or P4 in furin cleavage sites significantly affect cleavage efficiency in P. pastoris. Kex2 protease, but not YPS1, in P. pastoris is responsible for processing furin cleavage sites. Heterologous expression of furin or overexpression of Kex2 in P. pastoris effectively increased cleavage efficiencies of furin cleavage sites. CONCLUSIONS: Our investigation on the processing of furin cleavage sites provides important information for recombinant expression of furin precursors in P. pastoris. Furin or Kex2 overexpressing strains may be good choices for expressing precursors processed by furin in P. pastoris. BioMed Central 2018-11-08 /pmc/articles/PMC6223083/ /pubmed/30409181 http://dx.doi.org/10.1186/s12934-018-1020-x Text en © The Author(s) 2018 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
Huang, Yide
Long, Yanyu
Li, Suhuan
Lin, Ting
Wu, Jingwen
Zhang, Yafei
Lin, Yao
Investigation on the processing and improving the cleavage efficiency of furin cleavage sites in Pichia pastoris
title Investigation on the processing and improving the cleavage efficiency of furin cleavage sites in Pichia pastoris
title_full Investigation on the processing and improving the cleavage efficiency of furin cleavage sites in Pichia pastoris
title_fullStr Investigation on the processing and improving the cleavage efficiency of furin cleavage sites in Pichia pastoris
title_full_unstemmed Investigation on the processing and improving the cleavage efficiency of furin cleavage sites in Pichia pastoris
title_short Investigation on the processing and improving the cleavage efficiency of furin cleavage sites in Pichia pastoris
title_sort investigation on the processing and improving the cleavage efficiency of furin cleavage sites in pichia pastoris
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6223083/
https://www.ncbi.nlm.nih.gov/pubmed/30409181
http://dx.doi.org/10.1186/s12934-018-1020-x
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