Endoplasmic reticulum phospholipid scramblase activity revealed after protein reconstitution into giant unilamellar vesicles containing a photostable lipid reporter

Abstract Transbilayer movement of phospholipids in biological membranes is mediated by a diverse set of lipid transporters. Among them are scramblases that facilitate a rapid bi-directional movement of lipids without metabolic energy input. Here, we established a new fluorescence microscopy-based as...

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Autores principales:	Patricia P. M. Mathiassen, Anant K. Menon, Thomas Günther Pomorski
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/f23f41219aa64d99a12758d6c0b3ce78
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spelling	oai:doaj.org-article:f23f41219aa64d99a12758d6c0b3ce782021-12-02T16:08:07ZEndoplasmic reticulum phospholipid scramblase activity revealed after protein reconstitution into giant unilamellar vesicles containing a photostable lipid reporter10.1038/s41598-021-93664-02045-2322https://doaj.org/article/f23f41219aa64d99a12758d6c0b3ce782021-07-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-93664-0https://doaj.org/toc/2045-2322Abstract Transbilayer movement of phospholipids in biological membranes is mediated by a diverse set of lipid transporters. Among them are scramblases that facilitate a rapid bi-directional movement of lipids without metabolic energy input. Here, we established a new fluorescence microscopy-based assay for detecting phospholipid scramblase activity of membrane proteins upon their reconstitution into giant unilamellar vesicles formed from proteoliposomes by electroformation. The assay is based on chemical bleaching of fluorescence of a photostable ATTO-dye labeled phospholipid with the membrane-impermeant reductant sodium dithionite. We demonstrate that this new methodology is suitable for the study of the scramblase activity of the yeast endoplasmic reticulum at single vesicle level.Patricia P. M. MathiassenAnant K. MenonThomas Günther PomorskiNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-14 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Patricia P. M. Mathiassen Anant K. Menon Thomas Günther Pomorski Endoplasmic reticulum phospholipid scramblase activity revealed after protein reconstitution into giant unilamellar vesicles containing a photostable lipid reporter
description	Abstract Transbilayer movement of phospholipids in biological membranes is mediated by a diverse set of lipid transporters. Among them are scramblases that facilitate a rapid bi-directional movement of lipids without metabolic energy input. Here, we established a new fluorescence microscopy-based assay for detecting phospholipid scramblase activity of membrane proteins upon their reconstitution into giant unilamellar vesicles formed from proteoliposomes by electroformation. The assay is based on chemical bleaching of fluorescence of a photostable ATTO-dye labeled phospholipid with the membrane-impermeant reductant sodium dithionite. We demonstrate that this new methodology is suitable for the study of the scramblase activity of the yeast endoplasmic reticulum at single vesicle level.
format	article
author	Patricia P. M. Mathiassen Anant K. Menon Thomas Günther Pomorski
author_facet	Patricia P. M. Mathiassen Anant K. Menon Thomas Günther Pomorski
author_sort	Patricia P. M. Mathiassen
title	Endoplasmic reticulum phospholipid scramblase activity revealed after protein reconstitution into giant unilamellar vesicles containing a photostable lipid reporter
title_short	Endoplasmic reticulum phospholipid scramblase activity revealed after protein reconstitution into giant unilamellar vesicles containing a photostable lipid reporter
title_full	Endoplasmic reticulum phospholipid scramblase activity revealed after protein reconstitution into giant unilamellar vesicles containing a photostable lipid reporter
title_fullStr	Endoplasmic reticulum phospholipid scramblase activity revealed after protein reconstitution into giant unilamellar vesicles containing a photostable lipid reporter
title_full_unstemmed	Endoplasmic reticulum phospholipid scramblase activity revealed after protein reconstitution into giant unilamellar vesicles containing a photostable lipid reporter
title_sort	endoplasmic reticulum phospholipid scramblase activity revealed after protein reconstitution into giant unilamellar vesicles containing a photostable lipid reporter
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/f23f41219aa64d99a12758d6c0b3ce78
work_keys_str_mv	AT patriciapmmathiassen endoplasmicreticulumphospholipidscramblaseactivityrevealedafterproteinreconstitutionintogiantunilamellarvesiclescontainingaphotostablelipidreporter AT anantkmenon endoplasmicreticulumphospholipidscramblaseactivityrevealedafterproteinreconstitutionintogiantunilamellarvesiclescontainingaphotostablelipidreporter AT thomasguntherpomorski endoplasmicreticulumphospholipidscramblaseactivityrevealedafterproteinreconstitutionintogiantunilamellarvesiclescontainingaphotostablelipidreporter
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Endoplasmic reticulum phospholipid scramblase activity revealed after protein reconstitution into giant unilamellar vesicles containing a photostable lipid reporter

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