Optimized protocol for the identification of lipid droplet proteomes using proximity labeling proteomics in cultured human cells

Summary: Lipid droplets are endoplasmic reticulum-derived neutral lipid storage organelles that play critical roles in cellular lipid and energy homeostasis. Here, we present a protocol for the identification of high-confidence lipid droplet proteomes in a cell culture model. This approach overcomes...

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Main Authors:	Clark W.H. Peterson, Kirandeep K. Deol, Milton To, James A. Olzmann
Format:	Article
Language:	English
Published:	Elsevier 2021-06-01
Series:	STAR Protocols
Subjects:	Cell Biology Cell culture Cell Membrane Cell separation/fractionation Cell-based Assays Proteomics
Online Access:	http://www.sciencedirect.com/science/article/pii/S2666166721002860

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spelling	doaj-b2c07c0ec1364c289400ecb1cdc62f522021-06-21T04:25:47ZengElsevierSTAR Protocols2666-16672021-06-0122100579Optimized protocol for the identification of lipid droplet proteomes using proximity labeling proteomics in cultured human cellsClark W.H. Peterson0Kirandeep K. Deol1Milton To2James A. Olzmann3Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Nutritional Sciences and Toxicology, University of California, Berkeley, Berkeley, CA 94720, USADepartment of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Nutritional Sciences and Toxicology, University of California, Berkeley, Berkeley, CA 94720, USADepartment of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Nutritional Sciences and Toxicology, University of California, Berkeley, Berkeley, CA 94720, USADepartment of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Nutritional Sciences and Toxicology, University of California, Berkeley, Berkeley, CA 94720, USA; Miller Institute for Basic Research in Science, University of California, Berkeley, Berkeley, CA 94720, USA; Chan Zuckerberg Biohub, San Francisco, CA 94158, USA; Corresponding authorSummary: Lipid droplets are endoplasmic reticulum-derived neutral lipid storage organelles that play critical roles in cellular lipid and energy homeostasis. Here, we present a protocol for the identification of high-confidence lipid droplet proteomes in a cell culture model. This approach overcomes limitations associated with standard biochemical fractionation techniques, employing an engineered ascorbate peroxidase (APEX2) to biotinylate endogenous lipid droplet proteins in living cells for subsequent purification and identification by proteomics.For complete details on the use and execution of this protocol, please refer to Bersuker et al. (2018).http://www.sciencedirect.com/science/article/pii/S2666166721002860Cell BiologyCell cultureCell MembraneCell separation/fractionationCell-based AssaysProteomics
collection	DOAJ
language	English
format	Article
sources	DOAJ
author	Clark W.H. Peterson Kirandeep K. Deol Milton To James A. Olzmann
spellingShingle	Clark W.H. Peterson Kirandeep K. Deol Milton To James A. Olzmann Optimized protocol for the identification of lipid droplet proteomes using proximity labeling proteomics in cultured human cells STAR Protocols Cell Biology Cell culture Cell Membrane Cell separation/fractionation Cell-based Assays Proteomics
author_facet	Clark W.H. Peterson Kirandeep K. Deol Milton To James A. Olzmann
author_sort	Clark W.H. Peterson
title	Optimized protocol for the identification of lipid droplet proteomes using proximity labeling proteomics in cultured human cells
title_short	Optimized protocol for the identification of lipid droplet proteomes using proximity labeling proteomics in cultured human cells
title_full	Optimized protocol for the identification of lipid droplet proteomes using proximity labeling proteomics in cultured human cells
title_fullStr	Optimized protocol for the identification of lipid droplet proteomes using proximity labeling proteomics in cultured human cells
title_full_unstemmed	Optimized protocol for the identification of lipid droplet proteomes using proximity labeling proteomics in cultured human cells
title_sort	optimized protocol for the identification of lipid droplet proteomes using proximity labeling proteomics in cultured human cells
publisher	Elsevier
series	STAR Protocols
issn	2666-1667
publishDate	2021-06-01
description	Summary: Lipid droplets are endoplasmic reticulum-derived neutral lipid storage organelles that play critical roles in cellular lipid and energy homeostasis. Here, we present a protocol for the identification of high-confidence lipid droplet proteomes in a cell culture model. This approach overcomes limitations associated with standard biochemical fractionation techniques, employing an engineered ascorbate peroxidase (APEX2) to biotinylate endogenous lipid droplet proteins in living cells for subsequent purification and identification by proteomics.For complete details on the use and execution of this protocol, please refer to Bersuker et al. (2018).
topic	Cell Biology Cell culture Cell Membrane Cell separation/fractionation Cell-based Assays Proteomics
url	http://www.sciencedirect.com/science/article/pii/S2666166721002860
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Optimized protocol for the identification of lipid droplet proteomes using proximity labeling proteomics in cultured human cells

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