Figure 4. 16-AP Fluorescence Enhancement of Mutant 9, SlyD, and EcR1

Aliquots (1 mM) of Mutant 9, SlyD, or EcR1 was titrated with 16-AP and the fluorescence enhancement of 16-AP measured. The abscissa represents total 16-AP concentration added to the cuvette. Fluorescence was recorded with a photon counting fluorometer and the units on the ordinate represent photons counted per second (cps). Mutant 9 exhibited a significant fluorescence enhancement,while SlyD exhibited either no or only a slight fluorescence enhancement compared to buffer alone. Wild type EcR1 exhibited fluorescence enhancement as previously described with a binding capacity of roughly 0.5 ligands bound per molecule of protein [11,12], and the Mutant 9 binding curve may suggest one 16-AP molecule is bound per polypeptide.