LC-MS/MS Profiling of Protein Complexes

For LC-MS/MS profiling two biological replicates were used based on the high level of reproducibility between replicates. In previous studies most of the proteins had a reproducible peak between biological replicates (13 (link), 21 (link), 32 (link)). Ion exchange chromatography provided a high-resolution separation and 65 fractions were analyzed by mass spectrometry (analyzed on Sciex 5600 mass spectrometer). For the SEC and IEX profiling experiments that were analyzed to predict protein complex composition were analyzed on Sciex 5600 mass spectrometer. The SEC fractions that were analyzed to test for oligomerization changes in predicted AIMP1L-interactors by profiling the aimpl1 mutant were analyzed on Q Exactive mass spectrometer. For CoIP-MS pull downs three replicates were performed with antibodies against the protein of interest and negative controls and were analyzed on Q Exactive mass spectrometer.