Hello,
I know for sure protein x binds and modifies histones and am looking for identifying its cofactors that potentially involve in the binding and modifications. what is the best way to reach my gol?
Any help would be appreciated
Thanks
Sounds like a classical problem for Co-Immunoprecipitation, i.e use (magnetic) beads coupled to an e.g. anti-histone antibody or if possible your protein of interest, pull-down your protein ( or the histone - as a sort of internal control) and see what else gets precipitated. Mass spec should let you identify your interaction partners
Thank you so much Mr.Metzner. I am wondering how I can know the binding- proteins were recruited by protein X not by others if I will use anti- histone to do Co-IP. On the other hand, if I will use anti-protein X to do Co-IP, how can I know the binding proteins bound protein X with it was binding histones?
I appreciate your time
Ahmed
Dear Ahmed, the direct vs. indirect binding issue is critical with CoIP. Once you've got some candidates indentified, you'd have to confirm direct binding, i.e. via FRET
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