Fig. 2.

Experimental strategy to probe selective enrichment of PKA isoforms and their interacting partners using stable isotope labeling. Following the parallel affinity enrichments using C8 and C2 beads, proteins were digested in-solution with LysC and trypsin, respectively. The tryptic peptides originating from the pull-down using the C8 beads were chemically labeled using CD₂O (“heavy-label”) whereas those originating from C2 beads (and also C8_OCH₃ and EtOH beads) were labeled with CH₂O (“light-label”). Each set of CD₂O- and CH₂O-labeled samples were mixed in a 1:1 ratio (supplemental Fig. S1) and then analyzed by LC-coupled nanospray LTQ-FT-ICR mass spectrometry for protein identification and quantification.