Measuring in vivo protein turnover and exchange in yeast macromolecular assemblies

We present a comprehensive and robust protocol to track the dynamics of all proteins in a complex in yeast cells. A single member of the protein assembly is tagged and conditionally expressed, minimizing the perturbations to the protein complex. Then, SILAC labeling and affinity purification are use...

Descripción completa

Detalles Bibliográficos
Autores principales: Hakhverdyan, Zhanna, Molloy, Kelly R., Subbotin, Roman I., Fernandez-Martinez, Javier, Chait, Brian T., Rout, Michael P.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2021
Materias:
Protocol
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8433251/
https://www.ncbi.nlm.nih.gov/pubmed/34527957
http://dx.doi.org/10.1016/j.xpro.2021.100800
Descripción
Sumario:We present a comprehensive and robust protocol to track the dynamics of all proteins in a complex in yeast cells. A single member of the protein assembly is tagged and conditionally expressed, minimizing the perturbations to the protein complex. Then, SILAC labeling and affinity purification are used for the assessment of the whole protein complex dynamics. This method can determine and distinguish both subunit turnover and exchange specifically in an assembly to provide a comprehensive picture of assembly dynamics. For complete details on the use and execution of this protocol, please refer to Hakhverdyan et al. (2021).

Ejemplares similares