Immunoaffinity purification of endogenous proteins from S. cerevisiae for post-translational modification and protein interaction analysis

Abstract

Summary: Protein regulation by post-translational modifications and protein-protein interactions is critical to controlling molecular pathways. Here, we describe an immunoaffinity purification approach in Saccharomyces cerevisiae. The protocol uses an endogenously-expressed epitope-tagged protein and can be applied to the identification of post-translational modifications or protein binding partners. The lysine methyltransferase Set5 is used as an example here to purify phosphorylated Set5 and identify phosphosites; however, this approach can be applied to a diverse set of proteins in yeast.For complete details on the use and execution of this protocol, please refer to Jaiswal et al. (2020).

Keywords

• Model Organisms
• Signal Transduction
• Protein Biochemistry
• Proteomics
• Protein expression and purification