Lambda phage cro repressor interaction with its operator DNA: 2'-deoxy-5-fluorouracil OR3 analogues.

Abstract

The experiments here show that chemically synthesized DNA containing fluorine at selected sites can be used to test specific predictions of a model for cro repressor--operator interaction. This is done by observation of the perturbation to the fluorine-19 NMR spectra of analogues of OR3 synthesized with 2'-deoxy-5-fluorouracil at specific positions in the DNA helix. Although the three-dimensional structure of the cro repressor from phage lambda has been determined by Matthews and co-workers [Anderson, W., Ohlendorf, D., Takeda, Y., & Matthews, B. (1981) Nature (London) 290, 754-758], direct structural observations on the complex of the protein with its specific DNA recognition sequence, OR3, are limited. From that structure of the protein, alone, a model of its complex to DNA was built by fitting B-form DNA, with some distortion [Ohlendorf, D., Anderson, W., Fisher, R., Takeda, Y., & Matthews, B. (1982) Nature (London) 298, 718-723]. That model proposes that the cro repressor contacts only one side of this DNA double helix and a number of specific protein--DNA contacts. To test the model, 2'-deoxy-5-fluorouracil was used to place the fluorine-19 nuclear spin-label on the side of the DNA contacting the cro repressor and on the opposite side facing away from the cro repressor. The results presented here are consistent with the prediction that lambda phage cro repressor contacts only one side of the DNA double helix.