THE ALLOSTERIC INTERACTION BETWEEN D-GALACTOSE AND THE ESCHERICHIA-COLI GALACTOSE REPRESSOR PROTEIN

The Escherichia cell galactose repressor protein (GalR) inhibits trans cription of the gal operon upon binding to two operator sites (1-7).Th is DNA binding activity is inhibited when D-galactose or D-fucose bind s to GalR (8-14). Fluorescence spectroscopy was used to characterize t he single tryptophan of GalR and to investigate the interaction betwee n galactose and GalR. Fluorescence quenching experiments place both tr yptophan residues of the GalR dimer in similar, solvent-exposed locati ons. Galactose is shown to enhance the intrinsic tryptophan fluorescen ce of GalR, the source of which is not explained by a change in decay times, but is due to an increase in the pre exponential factor of the longest of the three fluorescence decay times. It is shown that the be ta-anomer of D-galactose is the likely form that binds to GalR. An inc rease in pH from 6.3 to 9.5 causes the equilibrium association constan t (K-a) describing the galactose-GalR interaction to decrease 10-fold. The interaction is cooperative below pH 9.5, Over the pH range of 6.3 to 9.5, the tryptophan solvent exposure of GalR increases. Galactose binding also induces an increase in exposure. These results, and other s presented in this paper, show that both pH and galactose cause globa l alterations in the structure of GalR.