Assaying the Posttranslational Arginylation of Proteins in Cultured Cells

• Published in: Methods in molecular biology (Clifton, N.J.) Vol. 1337; p. 49
• Main Authors: Galiano, Mauricio R, Hallak, Marta E
• Format: Journal Article
• Language: English
• Published: United States 2015
• Subjects: Aminoacyltransferases - metabolism; Animals; Arginine - metabolism; Cell Line; Cells, Cultured; Isotope Labeling; Protein Processing, Post-Translational; Proteins - chemistry; Proteins - metabolism; Sequence Analysis, Protein - methods; Staining and Labeling
• ISSN: 1940-6029
• DOI: 10.1007/978-1-4939-2935-1_7

To evaluate the posttranslational arginylation of proteins in vivo, we describe a protocol for studying the (14)C-Arg incorporation into proteins of cells in culture. The conditions determined for this particular modification contemplate both the biochemical requirements of the enzyme ATE1 and the adjustments that allowed the discrimination between posttranslational arginylation of proteins and de novo synthesis. These conditions are applicable for different cell lines or primary cultures, representing an optimal procedure for the identification and the validation of putative ATE1 substrates.