DNA loop formation between Nag repressor molecules bound to its two operator sites is necessary for repression of the nag regulon of Escherichia coli in vivo.

Binding sites for the Nag repressor overlap the transcription start sites of the divergent nagE and nagB genes, such that the centres of the sites are separated by nine turns of the B-DNA helix. Mutations which prevent repressor binding to either site or alter the phasing of the binding sites result in simultaneous derepression of both genes. An additional mutation which restores the phasing of the two sites permits repression. These observations show that repression is the result of co-operative binding of the repressor to its two sites, resulting in the formation of a loop of DNA.