The full-length mu-opioid receptor: a conformational study by circular dichroism in trifluoroethanol and membrane-mimetic environments.
The secondary structure content of the recombinant human mu-opioid receptor (HuMOR) solubilized in trifluoroethanol (TFE) and in detergent micelles was investigated by circular dichroism. In both conditions, this G protein-coupled receptor adopts a characteristic alpha-helical structure, with minima at 208 and 222 nm as observed in the circular dichroism spectra. After deconvolution of spectra, the alpha-helix contents were estimated to be in the range of 50% in TFE and in sodium dodecyl sulfate at pH 6. These values are in accordance with the predicted secondary structure content determined for the mu-opioid receptor. A pH-dependent effect was observed on the secondary structure of the receptor solubilized in detergents, which demonstrates the essential role of ionic and hydrophobic interactions on the secondary structure. Circular dichroism spectra of EGFP-HuMOR, a fusion protein between the enhanced green fluorescent protein (EGFP) and the mu-opioid receptor, and EGFP solubilized in TFE were also analyzed as part of this study.
Institut de Pharmacologie et de Biologie Structurale (UMR 5089), Centre National de la Recherche Scientifique, Université de Toulouse, 205 route de Narbonne, Toulouse Cedex 4, France.
Isabelle
I
Muller
Valérie
V
Sarramégna
Marie
M
Renault
Vincent
V
Lafaquière
Sarra
S
Sebai
Alain
A
Milon
Franck
F
Talmont
D002942:Circular Dichroism / Q000379:methods*; D003902:Detergents / Q000737:chemistry; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D057927:Hydrophobic and Hydrophilic Interactions; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D011869:Radioligand Assay; D017450:Receptors, Opioid, mu / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012995:Solubility; D014270:Trifluoroethanol / Q000737:chemistry*
D003902:Detergents; D008565:Membrane Proteins; D017450:Receptors, Opioid, mu; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D014270:Trifluoroethanol