title: Trifluoroethanol-induced conformational transition of the C-terminal sterile alpha motif (SAM) of human p73.
abstract: The alpha splice variant of p73 (p73α), a homologue of the tumour suppressor p53, has at its C terminus a sterile alpha motif (SAM); this domain, SAMp73, is involved in lipid binding and it is thought to mediate in protein-protein interactions. As SAMp73 is a 68-residue-long helical bundle, it could be a good model to study the (2,2,2-trifluoroethanol) TFE-induced conformational transitions of α-helical proteins. Furthermore, as SAMp73 binds to lipids through a well-known polypeptide patch, we can test whether TFE is a good mimic of lipids and membranes. To address those questions, we used several biophysical probes, namely, fluorescence, circular dichroism, 1D, 2D and 3D-NMR spectroscopies, and dynamic light scattering. The TFE-induced conformational transition of SAMp73 was complex, involving several species as detected by the biophysical probes. The last TFE-induced transition occurred at a concentration of TFE of ∼20% (v/v), where the protein lost its compactness. None of those TFE-induced species accumulated during the two-state folding of SAMp73 in aqueous solution. The final state at 40% TFE was highly helical, but its structure was not rigid. For SAMp73, TFE did not properly mimic a membrane-like environment, since at very low TFE concentrations, other residues, together with those known to interact with lipids, were also affected by the co-solvent. Comparison with studies on isolated peptides, comprising the helical regions of SAMp73, suggests that peptides were good models of the intact protein in TFE.
authors.affiliation: Instituto de Biología Molecular y Celular, Universidad Miguel Hernández, Elche, Alicante, Spain; Instituto de Biocomputación y Física de Sistemas Complejos, Zaragoza, Spain. Electronic address: jlneira@umh.es.
authors.forename: José L
authors.initials: JL
authors.lastname: Neira
authors.affiliation: Department of Chemistry and Physics, University of Almería, Agrifood Campus of International Excellence (ceiA3), Almería, Spain. Electronic address: acamara@ual.es.
authors.forename: Ana
authors.initials: A
authors.lastname: Cámara-Artigas
mesh_terms: D002462:Cell Membrane / Q000378:metabolism; D002942:Circular Dichroism; D006801:Humans; D008027:Light; D009682:Magnetic Resonance Spectroscopy; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D000072417:Protein Domains; D017510:Protein Folding; D025941:Protein Interaction Mapping; D017433:Protein Structure, Secondary; D012542:Scattering, Radiation; D012997:Solvents / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D000071682:Sterile Alpha Motif / Q000235:genetics*; D014270:Trifluoroethanol / Q000737:chemistry*; D000072160:Tumor Protein p73 / Q000737:chemistry*
chemical_list: D010455:Peptides; D012997:Solvents; C000606460:TP73 protein, human; D000072160:Tumor Protein p73; D014270:Trifluoroethanol
keywords: Circular dichroism; Fluorescence; Light-scattering; NMR; Protein conformation; TFE