Fig. 8.

Effect of the tmK72A mutation on the free energy landscape of the His73-heme mediated alkaline conformational transition of iso-1-Cytc. yWT/K73H has tmK72 and WT*/K73H carries the tmK72A mutation. The range given for the relative stabilities of the native conformer and His73-heme alkaline conformer for WT*/K73H is based on kinetic (0.7 kcal/mol; using k_f,His and k_b,His at pH 8 in Table 4 to obtained K_C1(His)) and thermodynamic data (0.9 kcal/mol, using pK_C1(His) in Table 2). The range in the magnitude of K_C1(His) produces a similar range for the relative stabilities of the His73-heme alkaline conformers of WT*/K73H and yWT/K73H. The change in the height of the barrier is calculated using the Eyring equation, yielding a decrease in the barrier for return to the native conformer by about 0.15 kcal/mol with the tmK72A mutation (k_b,His increases from 7 to 8.8 s⁻¹). The range in the ΔG of the TS for WT*/K73H versus yWT/K73H results from the range in the ΔG of the His73-heme conformers of WT*/K73H and yWT/K73H. Lower left: structure of native iso-1-Cytc (pdb code: 2YCC with Ω-loop D colored salmon shown as a space-filling model. Lower right: structure of the Lys73-heme alkaline conformer (pdb code: 1LMS) with Ω-loop D colored salmon. Met80, Tyr67, Pro71 and Ala/Lys72 are shown as space filling models. Ala72 in this structure has been converted to Lys using the mutate function of PyMol (added carbons are shown in gray).