Specific purine N7-nitrogens are critical for high affinity binding by the trp repressor.

We have analysed the interaction of the trp repressor with the trpEDCBA operator using a series of modified trp operator sequences incorporating two isosteric purine analogues that lack N7-nitrogens. Our results suggest that as well as the direct contact between Arg69 and G-9, three additional purine N7-nitrogens, implicated in specific, water-mediated contacts to the repressor, are critical for formation of the high-affinity repressor-operator complex. We conclude that the crystal structure obtained by Otwinowski et al. reflects high-affinity sequence-specific binding of the trp repressor to the trp operator, and that in some cases proteins can use water molecules to extend amino acid side chains in order to derive favorable binding energy in complex formation.