MUTAGENESIS OF AMINO-ACID-RESIDUES REQUIRED FOR BINDING OF COREPRESSORS TO THE PURINE REPRESSOR

The corepressor binding domain of the Escherichia coil purine represso r (PurR) is homologous with several periplasmic sugar-binding proteins . Four amino acids in PurR were investigated for a role in binding of corepressors. Three of the residues, Asp(146), Arg(196), and Asp(275) conserved in periplasmic binding proteins for ribose, glucose/galactos e, and arabinose and function to bind sugars. A fourth amino acid, Trp (147), required for corepressor binding to PurR, corresponds to residu es in glucose/galactose, ribose, and arabinose that also have a role i n sugar binding. The four mutations that were constructed perturbed th e binding of both hypoxanthine and guanine thus providing evidence for a single corepressor site/PurR subunit, The decreased corepressor bin ding affinity resulted in reduced affinity of mutant repressors for op erator DNA in vitro and decreased capacity for repression in vivo. The corepressor-binding site in PurR appears to be similar to the conserv ed ligand-binding sites in the three periplasmic sugar-binding protein s and in the LacI family of repressors.