Isolation of the gal repressor.

The repressor of the galactose operon of Escherichia coli has been partially purified and identified as a protein. Induction of a lysogen in which lambda was linked to the bacterial galR and lysine genes resulted in a large increase in the production of the gal repressor. Single-step purification by affinity chromatography, using the ligand p-aminophenyl-beta-D-thiogalactoside linked to beaded agarose, provided a convenient method of separating the gal repressor from other DNA-binding proteins. Binding of gal repressor to lambdapgal[(32)P]DNA was studied by assay of binding to a nitrocellulose filter. Interaction between gal repressor and lambdapgal DNA showed a high degree of specificity; the dissociation constant of the complex was estimated to be 1.0 x 10(-12) M. Unlabeled lambdapgal DNA competed for binding to gal repressor, but lambdaDNA and lambdah80dlac DNA did not. Fucose and galactose, which function as inducers of the galactose operon in vivo, produced one-half maximal inhibition of gal repressor-lambdapgal DNA binding at concentrations of 5 x 10(-5) M.