A highly efficient method for the production and purification of recombinant human CXCL8.

Chemokines play diverse and fundamental roles in the immune system and human disease, which has prompted their structural and functional characterisation. Production of recombinant chemokines that are folded and bioactive is vital to their study but is limited by the stringent requirements of a nati...

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Autores principales:	Sophie McKenna, Sean Patrick Giblin, Rosemarie Anne Bunn, Yingqi Xu, Stephen John Matthews, James Edward Pease
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/605d3d97aa614c1fb340f669ad356e21
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spelling	oai:doaj.org-article:605d3d97aa614c1fb340f669ad356e212021-12-02T20:13:39ZA highly efficient method for the production and purification of recombinant human CXCL8.1932-620310.1371/journal.pone.0258270https://doaj.org/article/605d3d97aa614c1fb340f669ad356e212021-01-01T00:00:00Zhttps://doi.org/10.1371/journal.pone.0258270https://doaj.org/toc/1932-6203Chemokines play diverse and fundamental roles in the immune system and human disease, which has prompted their structural and functional characterisation. Production of recombinant chemokines that are folded and bioactive is vital to their study but is limited by the stringent requirements of a native N-terminus for receptor activation and correct disulphide bonding required to stabilise the chemokine fold. Even when expressed as fusion proteins, overexpression of chemokines in E. coli tends to result in the formation of inclusion bodies, generating the additional steps of solubilisation and refolding. Here we present a novel method for producing soluble chemokines in relatively large amounts via a simple two-step purification procedure with no requirements for refolding. CXCL8 produced by this method has the correct chemokine fold as determined by NMR spectroscopy and in chemotaxis assays was indistinguishable from commercially available chemokines. We believe that this protocol significantly streamlines the generation of recombinant chemokines.Sophie McKennaSean Patrick GiblinRosemarie Anne BunnYingqi XuStephen John MatthewsJames Edward PeasePublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 16, Iss 10, p e0258270 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Sophie McKenna Sean Patrick Giblin Rosemarie Anne Bunn Yingqi Xu Stephen John Matthews James Edward Pease A highly efficient method for the production and purification of recombinant human CXCL8.
description	Chemokines play diverse and fundamental roles in the immune system and human disease, which has prompted their structural and functional characterisation. Production of recombinant chemokines that are folded and bioactive is vital to their study but is limited by the stringent requirements of a native N-terminus for receptor activation and correct disulphide bonding required to stabilise the chemokine fold. Even when expressed as fusion proteins, overexpression of chemokines in E. coli tends to result in the formation of inclusion bodies, generating the additional steps of solubilisation and refolding. Here we present a novel method for producing soluble chemokines in relatively large amounts via a simple two-step purification procedure with no requirements for refolding. CXCL8 produced by this method has the correct chemokine fold as determined by NMR spectroscopy and in chemotaxis assays was indistinguishable from commercially available chemokines. We believe that this protocol significantly streamlines the generation of recombinant chemokines.
format	article
author	Sophie McKenna Sean Patrick Giblin Rosemarie Anne Bunn Yingqi Xu Stephen John Matthews James Edward Pease
author_facet	Sophie McKenna Sean Patrick Giblin Rosemarie Anne Bunn Yingqi Xu Stephen John Matthews James Edward Pease
author_sort	Sophie McKenna
title	A highly efficient method for the production and purification of recombinant human CXCL8.
title_short	A highly efficient method for the production and purification of recombinant human CXCL8.
title_full	A highly efficient method for the production and purification of recombinant human CXCL8.
title_fullStr	A highly efficient method for the production and purification of recombinant human CXCL8.
title_full_unstemmed	A highly efficient method for the production and purification of recombinant human CXCL8.
title_sort	highly efficient method for the production and purification of recombinant human cxcl8.
publisher	Public Library of Science (PLoS)
publishDate	2021
url	https://doaj.org/article/605d3d97aa614c1fb340f669ad356e21
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A highly efficient method for the production and purification of recombinant human CXCL8.

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