Production of unstable proteins through the formation of stable core complexes

Nat Commun. 2016 Mar 17;7:10932. doi: 10.1038/ncomms10932.

Abstract

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

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

Substances

  • Multiprotein Complexes
  • Receptors, Cytoplasmic and Nuclear
  • Solvents