6 research outputs found
Apparent thermodynamic parameters for the equilibrium unfolding of RaPrP<sup>C</sup>(121–228) and the I214V mutant at 25°C.
<p>Note: is an estimate of the free energy in the absence of denaturant, the parameter represents the cooperativity of the unfolding transition, and is the concentration of urea at the midpoint of unfolding. The determined parameters for the wild-type <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0013273#pone.0013273-Wen1" target="_blank">[30]</a> are listed here to facilitate comparison.</p
Relaxation rates R<sub>1</sub>, R<sub>2</sub> and {<sup>1</sup>H}-<sup>15</sup>N heteronuclear NOEs of the I214V mutant of RaPrP<sup>C</sup>(121–228).
<p>Regular secondary structure elements are indicated on the top. The relaxation constants and the experimental errors were extracted by a single exponential curve fitting of the peak heights using Sparky (T. D. Goddard and D. G. Kneller, University of California, San Francisco).</p
Modelfree analysis.
<p>(A) Order parameters S<sup>2</sup> of the I214V mutant of RaPrP<sup>C</sup>(121–228). Regular secondary structure elements are indicated on the top. The program Fastmodelfree <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0013273#pone.0013273-Cole1" target="_blank">[63]</a> was used to perform the calculation. Unavailable S<sup>2</sup> values for a few residues are due to the absence of data or failure in the data fitting. (B) Differences in S<sup>2</sup> values between the wild-type and the mutant. The difference is calculated according to the equation: ΔS<sup>2</sup> = S<sup>2</sup><sub>mutant</sub>−S<sup>2</sup><sub>wild-type</sub>. The absence of ΔS<sup>2</sup> values for residues result from unavailable S<sup>2</sup> values for either the wild-type or the mutant. (C) ΔS<sup>2</sup> values are mapped onto the tertiary structures of the I214V mutant: blue for ΔS<sup>2</sup>≥0, red for ΔS<sup>2</sup><0, and grey for ΔS<sup>2</sup> unavailable. This ribbon diagram is generated by PyMol (kindly provided by Prof. DeLano WL).</p
Urea-induced transitions of the I214V mutant of RaPrP<sup>C</sup>(121–228) characterized by CD spectroscopy.
<p>(A) The folded protein in the absence of urea (solid line) compared with the unfolded protein in the presence of 9 M urea (dashed line). (B) Mean residue ellipticity measured at 222 nm (θ<sub>222</sub>) in the presence of urea of different concentration. θ<sub>222</sub> in denaturation is indicated by solid squares (▪), while in renaturation by open circles (○). The solid line presents the fitting curve on the basis of a two-state mechanism.</p
Solution structure of the I214V mutant of RaPrP<sup>C</sup>(91–228).
<p>(A) Diagram of 15 lowest-energy conformations. (B) Ribbon cartoon showing the secondary structure elements of the mean structure. (C) Sausage diagram showing the superposition of structures of the wild-type (grey) and the mutant (pink). The structure was determined in acetate buffer at pH 4.5. The diagrams are generated using MolMol <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0013273#pone.0013273-Koradi1" target="_blank">[61]</a>.</p
Reduced spectral density functions analysis.
<p>(A) Spectral densities of the I214V mutant of RaPrP<sup>C</sup>(121–228). (B) Differences of spectral densities between the wild-type and the I214V mutant. The difference is calculated as follows: Δ<i>J</i>(ω) = <i>J</i>(ω)<sub>mutant</sub>−<i>J</i>(ω)<sub>wild-type</sub>. Regular secondary structure elements are indicated on the top. The notebook provided by Spyracopoulos <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0013273#pone.0013273-Spyracopoulos1" target="_blank">[62]</a> was used to execute the calculation.</p