Phosphorylation Mapping of Atg13 in Response to Atg1 Activity and Rapamycin Treatment in Budding Yeast

We performed phosphorylation site mapping on of budding yeast Atg13 applying a quantitative mass spectrometry-based proteomics approach based on stable isotope labeling with amino acids in cell culture (SILAC). We determined changes in the Atg13 phosphorylation pattern in response of inhibition of kinase Atg1 and starvation conditions (using rapamycin), respectively. To do so, Atg13 was tandem purified via a histidine-biotin (HB) tag from cells growing at the respective experimental condition. Different stimulus-dependent PTM profiles of Atg13 have been identified.