Quantitative Assessment of the Effects of Trypsin Digestion Methods on Affinity Purification-Mass Spectrometry-based Protein-Protein Interaction Analysis.

ABSTRACT: Affinity purification-mass spectrometry (AP-MS) has become the method of choice for discovering protein-protein interactions (PPIs) under native conditions. The success of AP-MS depends on the efficiency of trypsin digestion and the recovery of the tryptic peptides for MS analysis. Several different protocols have been used for trypsin digestion of protein complexes in AP-MS studies, but no systematic studies have been conducted on the impact of trypsin digestion conditions on the identification of PPIs. Here, we used NFκB/RelA and Bromodomain-containing protein 4 (BRD4) as baits and test five distinct trypsin digestion methods (two using "on-beads," three using "elution-digestion" protocols). Although the performance of the trypsin digestion protocols change slightly depending on the diff

SUBMITTER: Zhang Y 

PROVIDER: S-EPMC5656008 | biostudies-literature | 2017 Aug

REPOSITORIES: biostudies-literature