Measuring in vivo protein turnover and exchange in yeast macromolecular assemblies

STAR Protoc. 2021 Sep 8;2(3):100800. doi: 10.1016/j.xpro.2021.100800. eCollection 2021 Sep 17.

Abstract

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).

Keywords: Protein Biochemistry; Proteomics.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Chromatography, Affinity / methods*
  • Isotope Labeling
  • Macromolecular Substances* / analysis
  • Macromolecular Substances* / chemistry
  • Macromolecular Substances* / metabolism
  • Proteolysis
  • Proteomics / methods*
  • Saccharomyces cerevisiae Proteins* / analysis
  • Saccharomyces cerevisiae Proteins* / chemistry
  • Saccharomyces cerevisiae Proteins* / metabolism

Substances

  • Macromolecular Substances
  • Saccharomyces cerevisiae Proteins