A tobacco etch virus protease with increased substrate tolerance at the P1' position.

Site-specific proteases are important tools for in vitro and in vivo cleavage of proteins. They are widely used for diverse applications, like protein purification, assessment of protein-protein interactions or regulation of protein localization, abundance or activity. Here, we report the developmen...

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Autores principales:	Christian Renicke, Roberta Spadaccini, Christof Taxis
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2013
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/a67faae0551041a9bf42215db8a28b86
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spelling	oai:doaj.org-article:a67faae0551041a9bf42215db8a28b862021-11-18T07:40:13ZA tobacco etch virus protease with increased substrate tolerance at the P1' position.1932-620310.1371/journal.pone.0067915https://doaj.org/article/a67faae0551041a9bf42215db8a28b862013-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/23826349/pdf/?tool=EBIhttps://doaj.org/toc/1932-6203Site-specific proteases are important tools for in vitro and in vivo cleavage of proteins. They are widely used for diverse applications, like protein purification, assessment of protein-protein interactions or regulation of protein localization, abundance or activity. Here, we report the development of a procedure to select protease variants with altered specificity based on the well-established Saccharomyces cerevisiae adenine auxotrophy-dependent red/white colony assay. We applied this method on the tobacco etch virus (TEV) protease to obtain a protease variant with altered substrate specificity at the P1' Position. In vivo experiments with tester substrates showed that the mutated TEV protease still efficiently recognizes the sequence ENLYFQ, but has almost lost all bias for the amino acid at the P1' Position. Thus, we generated a site-specific protease for synthetic approaches requiring in vivo generation of proteins or peptides with a specific N-terminal amino acid.Christian RenickeRoberta SpadacciniChristof TaxisPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 8, Iss 6, p e67915 (2013)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Christian Renicke Roberta Spadaccini Christof Taxis A tobacco etch virus protease with increased substrate tolerance at the P1' position.
description	Site-specific proteases are important tools for in vitro and in vivo cleavage of proteins. They are widely used for diverse applications, like protein purification, assessment of protein-protein interactions or regulation of protein localization, abundance or activity. Here, we report the development of a procedure to select protease variants with altered specificity based on the well-established Saccharomyces cerevisiae adenine auxotrophy-dependent red/white colony assay. We applied this method on the tobacco etch virus (TEV) protease to obtain a protease variant with altered substrate specificity at the P1' Position. In vivo experiments with tester substrates showed that the mutated TEV protease still efficiently recognizes the sequence ENLYFQ, but has almost lost all bias for the amino acid at the P1' Position. Thus, we generated a site-specific protease for synthetic approaches requiring in vivo generation of proteins or peptides with a specific N-terminal amino acid.
format	article
author	Christian Renicke Roberta Spadaccini Christof Taxis
author_facet	Christian Renicke Roberta Spadaccini Christof Taxis
author_sort	Christian Renicke
title	A tobacco etch virus protease with increased substrate tolerance at the P1' position.
title_short	A tobacco etch virus protease with increased substrate tolerance at the P1' position.
title_full	A tobacco etch virus protease with increased substrate tolerance at the P1' position.
title_fullStr	A tobacco etch virus protease with increased substrate tolerance at the P1' position.
title_full_unstemmed	A tobacco etch virus protease with increased substrate tolerance at the P1' position.
title_sort	tobacco etch virus protease with increased substrate tolerance at the p1' position.
publisher	Public Library of Science (PLoS)
publishDate	2013
url	https://doaj.org/article/a67faae0551041a9bf42215db8a28b86
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A tobacco etch virus protease with increased substrate tolerance at the P1' position.

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