Interaction of indoleacrylic acid with Trp aporepressor from Escherichia coli.
Trans-beta-Indoleacrylic acid (IAA) binds with moderately high affinity to the dimeric protein, trp aporepressor from Escherichia coli. IAA is itself nonfluorescent, and, upon binding, IAA quenches approximately 95% of the intrinsic tryptophan fluorescence of the protein. Since there is an overlap between the absorbance of IAA and the emission of tryptophan, this quenching appears to be due to resonance energy transfer. To adequately fit binding isotherms, it was necessary to use a model in which the binding of IAA to one subunit is able to quench fluorescence from both subunits of the protein. This model also assumes that binding to the two subunits occurs independently. Binding data were obtained as a function of the concentration of trp aporepressor, over the range of 1 x 10(-8) to 5 x 10(-5) M, in order to test the possibility that protein subunit association or dissociation occurs. When analyzed in terms of the above model, no significant protein concentration dependence is seen for the IAA association constant, indicating that either (i) the protein does not undergo changes in oligomerization over this concentration range or that (ii) different states of aggregation bind IAA with similar affinity. The affinity of IAA was found to increase at higher ionic strength and at lower pH. Also, evidence is presented for a photoinduced change in the configuration of IAA.
Department of Chemistry, University of Mississippi, University 38677.
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D001426:Bacterial Proteins; D004926:Escherichia coli; D006863:Hydrogen-Ion Concentration; D007211:Indoles / Q000378:metabolism*; D009994:Osmolar Concentration; D011485:Protein Binding; D011487:Protein Conformation; D012097:Repressor Proteins / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013057:Spectrum Analysis; D013816:Thermodynamics; D029968:Escherichia coli Proteins
D001426:Bacterial Proteins; D007211:Indoles; D012097:Repressor Proteins; C027190:TRPR protein, E coli; D029968:Escherichia coli Proteins; C001446:indoleacrylic acid