Development of Series of Affinity Tags in Streptomyces

Abstract Streptomyces are of great biological and industrial significance due to their complex morphological development and ability to produce numerous secondary metabolites. However, the intrinsic biochemical mechanisms underlying morphogenesis and secondary metabolism are rarely revealed, partial...

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Autores principales:	Xu-Ming Mao, Ning Sun, Yang Zheng, Yong-Quan Li
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2017
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/897abb4d8c714bddba416d1db1c902b2
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spelling	oai:doaj.org-article:897abb4d8c714bddba416d1db1c902b22021-12-02T12:32:55ZDevelopment of Series of Affinity Tags in Streptomyces10.1038/s41598-017-07377-42045-2322https://doaj.org/article/897abb4d8c714bddba416d1db1c902b22017-07-01T00:00:00Zhttps://doi.org/10.1038/s41598-017-07377-4https://doaj.org/toc/2045-2322Abstract Streptomyces are of great biological and industrial significance due to their complex morphological development and ability to produce numerous secondary metabolites. However, the intrinsic biochemical mechanisms underlying morphogenesis and secondary metabolism are rarely revealed, partially because of the limited availability of the biochemical tools in Streptomyces. Here we provided series of integrative vectors with various affinity tags, including single tags 3×FLAG, 3×HA, 3×Strep-tag II, 18×His, 13×Myc, and dual tags, all of which were driven from a strong constitutive promoter ermEp*. Using a sigma factor SigT from S. coelicolor as a model, we successfully expressed and immuno-detected SigT fused with all tags. Moreover, after SigT was N-terminally tagged with 3×FLAG and C-terminally tagged with 18×His, we isolated SigT-interactive proteins from the S. coelicolor lysate based on the tandem affinity purification (TAP). Particularly, among the proteins purified, the SigT cognate anti-sigma factor RstA ranked the top with the most total independent spectra. These data suggested the feasibility of these affinity tags in Streptomyces, which will be widely employed to explore the biochemical mechanisms to further understand the dynamic and elaborate regulation in this genus.Xu-Ming MaoNing SunYang ZhengYong-Quan LiNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 7, Iss 1, Pp 1-7 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Xu-Ming Mao Ning Sun Yang Zheng Yong-Quan Li Development of Series of Affinity Tags in Streptomyces
description	Abstract Streptomyces are of great biological and industrial significance due to their complex morphological development and ability to produce numerous secondary metabolites. However, the intrinsic biochemical mechanisms underlying morphogenesis and secondary metabolism are rarely revealed, partially because of the limited availability of the biochemical tools in Streptomyces. Here we provided series of integrative vectors with various affinity tags, including single tags 3×FLAG, 3×HA, 3×Strep-tag II, 18×His, 13×Myc, and dual tags, all of which were driven from a strong constitutive promoter ermEp*. Using a sigma factor SigT from S. coelicolor as a model, we successfully expressed and immuno-detected SigT fused with all tags. Moreover, after SigT was N-terminally tagged with 3×FLAG and C-terminally tagged with 18×His, we isolated SigT-interactive proteins from the S. coelicolor lysate based on the tandem affinity purification (TAP). Particularly, among the proteins purified, the SigT cognate anti-sigma factor RstA ranked the top with the most total independent spectra. These data suggested the feasibility of these affinity tags in Streptomyces, which will be widely employed to explore the biochemical mechanisms to further understand the dynamic and elaborate regulation in this genus.
format	article
author	Xu-Ming Mao Ning Sun Yang Zheng Yong-Quan Li
author_facet	Xu-Ming Mao Ning Sun Yang Zheng Yong-Quan Li
author_sort	Xu-Ming Mao
title	Development of Series of Affinity Tags in Streptomyces
title_short	Development of Series of Affinity Tags in Streptomyces
title_full	Development of Series of Affinity Tags in Streptomyces
title_fullStr	Development of Series of Affinity Tags in Streptomyces
title_full_unstemmed	Development of Series of Affinity Tags in Streptomyces
title_sort	development of series of affinity tags in streptomyces
publisher	Nature Portfolio
publishDate	2017
url	https://doaj.org/article/897abb4d8c714bddba416d1db1c902b2
work_keys_str_mv	AT xumingmao developmentofseriesofaffinitytagsinstreptomyces AT ningsun developmentofseriesofaffinitytagsinstreptomyces AT yangzheng developmentofseriesofaffinitytagsinstreptomyces AT yongquanli developmentofseriesofaffinitytagsinstreptomyces
_version_	1718393931090100224

Development of Series of Affinity Tags in Streptomyces

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