Production of unstable proteins through the formation of stable core complexes.

Date publication

mars 2016

Journal

Nature communications

Auteurs

Membres identifiés du Cancéropôle Est :
Dr RUFF Marc

Tous les auteurs :
Levy N, Eiler S, Pradeau-Aubreton K, Maillot B, Stricher F, Ruff M

Lien Pubmed

https://www.ncbi.nlm.nih.gov/pubmed/26983699

Résumé

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.

Mots clés

Cell Line, HIV-1, metabolism, Humans, Multiprotein Complexes, isolation & purification, Protein Stability, Receptors, Cytoplasmic and Nuclear, metabolism, Solubility, Solvents

Référence

Nat Commun. 2016 Mar;7:10932