Production of unstable proteins through the formation of stable core complexes

Nicolas Levy (1) , Sylvia Eiler (1) , Karine Pradeau-Aubreton (1) , Benoit Maillot (1) , François Stricher (1) , Marc Ruff (1)

1 IGBMC - Institut de Génétique et de Biologie Moléculaire et Cellulaire

Purification of proteins that participate in large transient complexes is impeded by low amounts, heterogeneity, instability and poor solubility. To circumvent these difficulties we set up a methodology that enables the production of stable complexes for structural and functional studies. This procedure is benchmarked and applied to two challenging protein families: the human steroid nuclear receptors (SNR) and the HIV-1 pre-integration complex. In the context of transcriptional regulation studies, we produce and characterize the ligand-binding domains of the glucocorticoid nuclear receptor and the oestrogen receptor beta in complex with a TIF2 (transcriptional intermediary factor 2) domain containing the three SNR-binding motifs. In the context of retroviral integration, we demonstrate the stabilization of the HIV-1 integrase by formation of complexes with partner proteins and DNA. This procedure provides a powerful research tool for structural and functional studies of proteins participating in non-covalent macromolecular complexes.

Domaines: Biochimie [q-bio.BM]; Biologie structurale [q-bio.BM]

Nicolas Levy, Sylvia Eiler, Karine Pradeau-Aubreton, Benoit Maillot, François Stricher, et al.. Production of unstable proteins through the formation of stable core complexes. Nature Communications, 2016, 7, ⟨10.1038/ncomms10932⟩. ⟨hal-03531830⟩