Coimmunoprecipitation is a powerful and commonly used method to identify protein–protein interactions
in a physiological context. Here, we report a coimmunoprecipitation protocol that was adapted and
optimized for the haloarchaeon Haloferax volcanii to identify interacting partners to the LonB protease.
This protocol includes the in vivo cross-linking of H. volcanii proteins using two different crosslinker
agents, dithiobis(succinimidyl propionate) and formaldehyde, followed by immunoprecipitation with anti-
LonB antibody conjugated to Protein A - Sepharose beads. Tryptic on-bead protein digestion was
performed combined with Mass Spectrometry analysis of peptides for the identification and quantification
of LonB ligands.
Paggi, R.A., De Castro, R.E., Cerletti, M. (2022). In Vivo Protein Cross-Linking and Coimmunoprecipitation in Haloferax volcanii. In: Ferreira-Cerca, S. (eds) Archaea. Methods in Molecular Biology, vol 2522. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2445-6_19
Link to chapter: https://link.springer.com/protocol/10.1007/978-1-0716-2445-6_19