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Figure 6. Interaction of nucleotides with residue V634 in the ATP-binding site explored by double mutant cycle analyses. (A) Mutant cycle analysis of the interaction of the nucleotide with residue V634. The ATP and CTP dose–response curves of WT CLC-1 and the V634F mutant were first constructed using the parameter Poc(ATP)/Poc(control) as the dependent parameter. The values of Ko and Kc (Table III) of ATP/CTP modulations were determined by fitting the data points to Eq. 2, and the Kc values were used to calculate the free energy differences of single or double manipulations (bar graph) according to Eq. 3 and the coupling constant, Ω, according to Eq. 4. n = 6–8. (B) Mutant cycle analysis of the nucleotide interaction with V634 using ATP and dATP. The ligand dose–response curves of WT CLC-1 and V634F were constructed using Poc(ATP)/Poc(control) as the dependent parameter. Ko and Kc were determined (Table III), and the Ω value and the free energy differences were calculated accordingly. n = 4–8. Image published in: Tseng PY et al. (2011) © 2011 Tseng et al. Creative Commons Attribution-NonCommercial-ShareAlike license Larger Image Printer Friendly View

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