Hydrogen exchange of the tetramerization domain of the human tumour suppressor p53 probed by denaturants and temperature.
We have analysed the hydrogen/deuterium exchange of the tetramerization domain of human tumour suppressor p53 under mild chemical denaturation conditions, and at different temperatures. Exchange behaviour has been measured for 16 amide protons in the chemical-denaturation studies and for seven protons in the temperature-denaturation studies. The exchange rates are in the range observed for other proteins with similar elements of secondary structure. The slowest-exchange core includes contributions from residues in the alpha helix and the beta sheet. However, only some of the slowest-exchanging protons correspond to residues involved in native interactions in the transient intermediate detected during the folding of this domain. The guanidinium-chloride denaturation curves of all residues seem to merge together, although they are well below the main isotherm of global unfolding. Thus, there is no evidence for several subglobal unfolding units. The activation parameters obtained from the temperature-denaturation experiments are similar to those obtained for monomeric proteins, and well below the global unfolding enthalpy obtained by circular dichroism measurements. Thus, the exchange studies at different denaturant concentrations and temperatures indicate that no particular folding intermediate is populated under those conditions.
Neira
J L
JL
Centro de Biología Molecular y Celular, Universidad Miguel Hernández, Alicante, Spain. jlneira@umh.es
Mateu
M G
MG
D019281:Dimerization; D019791:Guanidine / Q000494:pharmacology; D006801:Humans; D006859:Hydrogen / Q000378:metabolism*; D006860:Hydrogen Bonding; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D011489:Protein Denaturation / Q000187:drug effects; D017510:Protein Folding; D020836:Protein Structure, Quaternary / Q000187:drug effects; D017434:Protein Structure, Tertiary / Q000187:drug effects; D013696:Temperature; D013816:Thermodynamics; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry* / Q000378:metabolism*
D016159:Tumor Suppressor Protein p53; D006859:Hydrogen; D019791:Guanidine