Figure 1

A typical AP-MS workflow. A typical AP-MS study consists of performing a set of experiments on bait proteins of interest, with the goal of identifying their interaction partners. In each experiment, a bait protein is tagged (e.g., using a FLAG-tag or TAP-tag) and expressed in cells. The bait protein and interacting prey proteins are affinity purified. The resulting mixture of bait and bound prey proteins is trypsinized into peptide fragments, which are separated by liquid chromatography and passed to a mass spectrometer for analysis. The mass spectrometer produces intensity spectra, which are matched to peptides to deduce proteins present in the purification. Interacting preys thus identified are assigned semi-quantitative spectral counts (SpC) indicating the propensity of each prey to bind to the bait. Data is collated from across the experiments into a matrix of bait-prey spectral counts, which serves as the input to post-processing methods that filter contaminants and identify true interactions.