answer 6-2: fit the signal with the equation for single site protein
Congratulations, this is quite a good answer! Unfortunately, as we shall see, it does not always work.
The equation you use is the same we derived in tutorial #1, i.e.:
You proceed as indicated and you obtain: Kd = 1 mM, offset = 0.2, Δ signal = 0.4. The residuals are good and you estimate that you recorded approximately the 0% - 75% ligand saturation range:
This method only yields acceptable results if the highest ligand concentration you could reach produced [PX]/[P]tot significantly higher than 50%; i.e. if the solubility of the ligand exceeds Kd by at least 3- to 5-folds.
If this is not the case, and the ligand's solubility equals or is lower than Kd, this method yields unreliable estimates of the fraction of liganded sites and Kd. Proceed to the next page for further discussion.
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