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Structural biology workflow for the expression and characterization of functional human sodium glucose transporter type 1 in Pichia pastoris

dc.contributor.authorSuades, Albert
dc.contributor.authorAlcaraz, Antonio
dc.contributor.authorCruz, Esteban
dc.contributor.authorÁlvarez-Marimon, Elena
dc.contributor.authorWhitelegge, Julian
dc.contributor.authorManyosa, Joan
dc.contributor.authorCladera, Josep
dc.contributor.authorPerálvarez-Marín, Alex
dc.date.accessioned2019-05-15T17:56:07Z
dc.date.available2019-05-15T17:56:07Z
dc.date.issued2019
dc.identifier.citationSuades, Albert, et al. "Structural biology workflow for the expression and characterization of functional human sodium glucose transporter type 1 in Pichia pastoris." Scientific reports, 2019, vol. 9, núm. 1ca_CA
dc.identifier.issn2045-2322
dc.identifier.urihttp://hdl.handle.net/10234/182498
dc.description.abstractHeterologous expression of human membrane proteins is a challenge in structural biology towards drug discovery. Here we report a complete expression and purification process of a functional human sodium/D-glucose co-transporter 1 (hSGLT1) in Pichia pastoris as representative example of a useful strategy for any human membrane protein. hSGLT1 gene was cloned in two different plasmids to develop parallel strategies: one which includes green fluorescent protein fusion for screening optimal conditions, and another for large scale protein production for structural biology and biophysics studies. Our strategy yields at least 1 mg of monodisperse purified recombinant hSGLT1 per liter of culture, which can be characterized by circular dichroism and infrared spectroscopy as an alpha-helical fold protein. This purified hSGLT1 transports co-substrates (Na+ and glucose) and it is inhibited by phlorizin in electrophysiological experiments performed in planar lipid membranes.ca_CA
dc.format.extent11 p.ca_CA
dc.format.mimetypeapplication/pdfca_CA
dc.language.isoengca_CA
dc.publisherNature Researchca_CA
dc.relation.isPartOfScientific reports, 2019, vol. 9, núm. 1ca_CA
dc.rightsOpen Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Cre-ative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not per-mitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.© The Author(s) 2019ca_CA
dc.rightsAtribución 4.0 Internacional*
dc.rights.urihttp://creativecommons.org/licenses/by-sa/4.0/*
dc.titleStructural biology workflow for the expression and characterization of functional human sodium glucose transporter type 1 in Pichia pastorisca_CA
dc.typeinfo:eu-repo/semantics/articleca_CA
dc.identifier.doihttps://doi.org/10.1038/s41598-018-37445-2
dc.relation.projectIDThis work was supported by the MICINN grants BFU2012-40137-C02-01 to J.C. and SAF2010-21385 to A.P.-M., and the NIH grant NIH P30 DK063491 to J.P.W.; A.P.-M. was a recipient of a Marie Curie International Outgoing Fellowship within the 7th European Community Framework Programme (PIOF-GA-2009-237120 to A.P.-M.) and a Universitat Autònoma de Barcelona-Programa Banco de Santander Fellowship. A.S. was a recipient of a Ph.D. fellowship and a travel bursary from Universitat Autònoma de Barcelonaca_CA
dc.rights.accessRightsinfo:eu-repo/semantics/openAccessca_CA
dc.relation.publisherVersionhttps://www.nature.com/articles/s41598-018-37445-2ca_CA
dc.type.versioninfo:eu-repo/semantics/publishedVersionca_CA


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Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Cre-ative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not per-mitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.© The Author(s) 2019
Except where otherwise noted, this item's license is described as Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Cre-ative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not per-mitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.© The Author(s) 2019