Artículo
Determination of the lipid composition of the GPI anchor
Autor/es | Aguilera Romero, María Auxiliadora
Sabido Bozo, Susana López Martín, Sergio Cortés Gómez, Alejandro Rodríguez Gallardo, Sofía Pérez Linero, Ana María Riezman, Isabelle Riezman, Howard Muñiz Guinea, Manuel |
Departamento | Universidad de Sevilla. Departamento de Biología Celular |
Fecha de publicación | 2021 |
Fecha de depósito | 2022-05-16 |
Publicado en |
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Resumen | In eukaryotic cells, a subset of cell surface proteins is attached by the glycolipid glycosylphosphatidylinositol (GPI) to the external leaflet of the plasma membrane where they play important roles as enzymes, receptors, ... In eukaryotic cells, a subset of cell surface proteins is attached by the glycolipid glycosylphosphatidylinositol (GPI) to the external leaflet of the plasma membrane where they play important roles as enzymes, receptors, or adhesion molecules. Here we present a protocol for purification and mass spectrometry analysis of the lipid moiety of individual GPI-anchored proteins (GPI-APs) in yeast. The method involves the expression of a specific GPI-AP tagged with GFP, solubilization, immunoprecipitation, separation by electrophoresis, blotting onto PVDF, release and extraction of the GPI-lipid moiety and analysis by mass spectrometry. By using this protocol, we could determine the precise GPI-lipid structure of the GPI-AP Gas1-GFP in a modified yeast strain. This protocol can be used to identify the lipid composition of the GPI anchor of distinct GPI-APs from yeast to mammals and can be adapted to determine other types of protein lipidation. |
Agencias financiadoras | Ministerio de Ciencia, Innovación y Universidades (MICINN). España European Commission (EC). Fondo Europeo de Desarrollo Regional (FEDER) Universidad de Sevilla Swiss National Science Foundation (SNFS) |
Identificador del proyecto | BFU2017-89700-P
VIPPIT-2020-I.5 51NF40-185898 310030_184949 |
Cita | Aguilera Romero, M.A., Sabido Bozo, S., López Martín, S., Cortés Gómez, A., Rodríguez Gallardo, S., Pérez Linero, A.M.,...,Muñiz Guinea, M. (2021). Determination of the lipid composition of the GPI anchor. PLOS ONE, 16 (8), e0256184. |
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10.1371_journal.pone.0256184.pdf | 606.6Kb | [PDF] | Ver/ | |