Authors:
H Mohammed, C Taylor, GD Brown, EK Papachristou, JS Carroll, CS D'Santos
Journal name: 
Nat Protoc
Citation info: 
11(2):316-326
Abstract: 
Rapid immunoprecipitation mass spectrometry of endogenous protein (RIME) is a method that allows the study of protein complexes, in particular chromatin and transcription factor complexes, in a rapid and robust manner by mass spectrometry (MS). The method can be used in parallel with chromatin immunoprecipitation-sequencing (ChIP-seq) experiments to provide information on both the cistrome and interactome for a given protein. The method uses formaldehyde fixation to stabilize protein complexes. By using antibodies against the endogenous target, the cross-linked complex is immunoprecipitated, rigorously washed, and then digested into peptides while avoiding antibody contamination (on-bead digestion). By using this method, MS identification of the target protein and several dozen interacting proteins is possible using a 100-min LC-MS/MS run. The protocol does not require substantial proteomics expertise, and it typically takes 2-3 d from the collection of material to results.
DOI: 
http://doi.org/10.1038/nprot.2016.020
Research group: 
Carroll Group
E-pub date: 
01 Feb 2016