Table 1 Thermodynamic parameters obtained from equilibrium unfolding experiments.

From: Global Implications of Local Unfolding Phenomena, Probed by Cysteine Reactivity in Human Frataxin

Variant

Urea*

Temperature†

Fluorescence

CD

CD

\({\boldsymbol{\Delta }}{\boldsymbol{\Delta }}{\boldsymbol{G}}{^\circ }_{{\bf{N}}\rightleftharpoons {\bf{U}}}^{{{\bf{H}}}_{{\bf{2}}}{\bf{O}}}\) (kcal mol−1)

\({\boldsymbol{\Delta }}{\boldsymbol{G}}{^\circ }_{{\bf{N}}\rightleftharpoons {\bf{U}}}^{{{\bf{H}}}_{{\bf{2}}}{\bf{O}}}\) (kcal mol−1)

Cm (M)

\({\boldsymbol{\Delta }}{\boldsymbol{\Delta }}{\boldsymbol{G}}{^\circ }_{{\bf{N}}\rightleftharpoons {\bf{U}}}^{{{\bf{H}}}_{{\bf{2}}}{\bf{O}}}\) (kcal mol−1)

\({\boldsymbol{\Delta }}{\boldsymbol{\Delta }}{\boldsymbol{G}}{^\circ }_{{\bf{N}}\rightleftharpoons {\bf{U}}}^{{{\bf{H}}}_{{\bf{2}}}{\bf{O}}}\) (kcal mol−1)

Cm (M)

Tm (°C)

FXN90–210

—

09.1 ± 0.5

4.94 ± 0.03

—

9.0 ± 0.5

4.92 ± 0.04

69.4 ± 0.4

FXN L198C

−1.0

08.1 ± 0.5

4.41 ± 0.03

90.9

8.1 ± 0.5

4.42 ± 0.05

66.1 ± 0.3

FXN L200C

−2.0

07.0 ± 0.4

3.85 ± 0.03

−1.8

7.2 ± 0.5

3.95 ± 0.04

62.3 ± 0.3

FXN L203C

−1.0

10.0 ± 0.6

5.47 ± 0.03

−0.8

9.8 ± 0.6

5.35 ± 0.06

71.0 ± 0.4

  1. *A two-state model was simultaneously fitted to the data obtained in urea-induced unfolding experiments followed by CD and Trp fluorescence for all variants (Fig. 2A,B). The value of the \({{\rm{m}}}_{{\rm{N}}\rightleftharpoons {\rm{U}}}^{}\) parameter was assumed the same for all variants, and found to be 1.8 ± 0.1 kcal mol−1 M−1, considerably larger than the value inferred by considering the protein’s length (1.5 kcal mol−1 M−1)33. \({\rm{\Delta }}{\rm{\Delta }}G{^\circ }_{{\rm{N}}\rightleftharpoons {\rm{U}}}^{{{\rm{H}}}_{{\rm{2}}}{\rm{O}}}={\rm{\Delta }}G{^\circ }_{{\rm{N}}\rightleftharpoons {\rm{U}}}^{{{\rm{H}}}_{{\rm{2}}}{\rm{O}},\,\mathrm{wt}}-{\rm{\Delta }}G{^\circ }_{{\rm{N}}\rightleftharpoons {\rm{U}}}^{{{\rm{H}}}_{{\rm{2}}}{\rm{O}},\,\mathrm{mutant}}\). Cm is the [urea] where 50% of the molecules are unfolded \(({\rm{\Delta }}G{{}^{\circ }}_{{\rm{N}}\rightleftharpoons {\rm{U}}}^{{{\rm{C}}}_{{\rm{m}}}^{}}=0)\).
  2. †A two-state model was simultaneously fitted to the data obtained in temperature-induced unfolding experiments followed by CD at 220 nm (Fig. 2C). The value of the \({{\rm{\Delta }}C}_{{{\rm{P}}}_{{\rm{N}}\rightleftharpoons {\rm{U}}}}\) parameter—the difference in the heat capacity between the native and unfolded states—was assumed the same for all variants and found to be 1.8 ± 0.3 kcal mol−1 K−1. Tm is the temperature where 50% of the molecules are unfolded \(({\rm{\Delta }}G{{}^{\circ }}_{{\rm{N}}\rightleftharpoons {\rm{U}}}^{{{\rm{T}}}_{{\rm{m}}}^{}}=0)\).
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