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Comparison of periplasmic and intracellular expression of Arabidopsis thionin proproteins in E. coli

Thionins are antimicrobial plant peptides produced as preproproteins consisting of a signal peptide, the thionin domain, and a so-called acidic domain. Only thionin itself has been isolated from plants. To study the processing of the precursor, it has to be produced in a heterologous system. Since b...

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Autores principales: Abbas, Amjad, Plattner, Stephan, Shah, Kausar Hussain, Bohlmann, Holger
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Springer Netherlands 2013
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3677976/
https://www.ncbi.nlm.nih.gov/pubmed/23515894
http://dx.doi.org/10.1007/s10529-013-1180-z
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author Abbas, Amjad
Plattner, Stephan
Shah, Kausar Hussain
Bohlmann, Holger
author_facet Abbas, Amjad
Plattner, Stephan
Shah, Kausar Hussain
Bohlmann, Holger
author_sort Abbas, Amjad
collection PubMed
description Thionins are antimicrobial plant peptides produced as preproproteins consisting of a signal peptide, the thionin domain, and a so-called acidic domain. Only thionin itself has been isolated from plants. To study the processing of the precursor, it has to be produced in a heterologous system. Since both domains contain several cysteines and, due to the known antimicrobial activity of the thionin, we tested the expression of all four Arabidopsis proproteins as fusion proteins. Periplasmic expression as fusion with maltose binding protein was not successful but cytoplasmic expression as His-tagged TRX fusion proteins with a TEV recognition sequence resulted in proteins of correct size. Use of the SHuffle strain C3030 further improved the expression. Fusion proteins inhibited growth of Escherichia coli. They could be cleaved by TEV protease, releasing authentic proproteins without any additional amino acid at the N-terminus. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s10529-013-1180-z) contains supplementary material, which is available to authorized users.
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spelling pubmed-36779762013-06-11 Comparison of periplasmic and intracellular expression of Arabidopsis thionin proproteins in E. coli Abbas, Amjad Plattner, Stephan Shah, Kausar Hussain Bohlmann, Holger Biotechnol Lett Original Research Paper Thionins are antimicrobial plant peptides produced as preproproteins consisting of a signal peptide, the thionin domain, and a so-called acidic domain. Only thionin itself has been isolated from plants. To study the processing of the precursor, it has to be produced in a heterologous system. Since both domains contain several cysteines and, due to the known antimicrobial activity of the thionin, we tested the expression of all four Arabidopsis proproteins as fusion proteins. Periplasmic expression as fusion with maltose binding protein was not successful but cytoplasmic expression as His-tagged TRX fusion proteins with a TEV recognition sequence resulted in proteins of correct size. Use of the SHuffle strain C3030 further improved the expression. Fusion proteins inhibited growth of Escherichia coli. They could be cleaved by TEV protease, releasing authentic proproteins without any additional amino acid at the N-terminus. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s10529-013-1180-z) contains supplementary material, which is available to authorized users. Springer Netherlands 2013-03-21 2013 /pmc/articles/PMC3677976/ /pubmed/23515894 http://dx.doi.org/10.1007/s10529-013-1180-z Text en © The Author(s) 2013 https://creativecommons.org/licenses/by/2.0/ Open AccessThis article is distributed under the terms of the Creative Commons Attribution License which permits any use, distribution, and reproduction in any medium, provided the original author(s) and the source are credited.
spellingShingle Original Research Paper
Abbas, Amjad
Plattner, Stephan
Shah, Kausar Hussain
Bohlmann, Holger
Comparison of periplasmic and intracellular expression of Arabidopsis thionin proproteins in E. coli
title Comparison of periplasmic and intracellular expression of Arabidopsis thionin proproteins in E. coli
title_full Comparison of periplasmic and intracellular expression of Arabidopsis thionin proproteins in E. coli
title_fullStr Comparison of periplasmic and intracellular expression of Arabidopsis thionin proproteins in E. coli
title_full_unstemmed Comparison of periplasmic and intracellular expression of Arabidopsis thionin proproteins in E. coli
title_short Comparison of periplasmic and intracellular expression of Arabidopsis thionin proproteins in E. coli
title_sort comparison of periplasmic and intracellular expression of arabidopsis thionin proproteins in e. coli
topic Original Research Paper
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3677976/
https://www.ncbi.nlm.nih.gov/pubmed/23515894
http://dx.doi.org/10.1007/s10529-013-1180-z
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