Affinity Purification of Glycosylphosphatidylinositol-anchored Proteins by Alpha-Toxin.
Methods Mol Biol
; 2303: 251-257, 2022.
Article
en En
| MEDLINE
| ID: mdl-34626384
ABSTRACT
The glycosylphosphatidylinositol (GPI)-anchor modification attaches a lipid anchor to the C-terminus of a protein, tethering the protein to the cell surface membrane. From this membrane-bound state, GPI-anchored proteins (GPI-APs) can be released into the extracellular space by multiple mechanisms, including proteolytic shedding and GPI lipase activity. Since the core GPI structure is co-released with the protein by GPI lipase activity, while removed from the protein by proteolytic cleavage, affinity purification by alpha-toxin (αToxin), which binds to the core domain of the GPI-anchor, isolates GPI-containing proteins from the culture medium. The following method details a technique for affinity purification of GP-APs using His-tagged αToxin for identification of GPI-anchored proteins, analysis of the GPI-anchor status of a protein of interest, or purification for subsequent biochemical analysis.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Cromatografía de Afinidad
Idioma:
En
Revista:
Methods Mol Biol
Asunto de la revista:
BIOLOGIA MOLECULAR
Año:
2022
Tipo del documento:
Article
País de afiliación:
Estados Unidos