The bacteriophage D108 Ner repressor binds a conformationally distinct operator.

The Ner protein encoded by the transposable coliphage D108, an 8.6 kDa lambda Cro-like repressor, binds to an operator spanning 50 bp of DNA. The distinguishing features of this operator are two perfect 11-bp inverted repeats (5'-CCGTGAGCTAC-3') that are separated by an 8-bp AT-rich spacer. Hyperreactivity of the ner operator to potassium permanganate and the hydroxyl radical indicate that the AT-rich spacer assumes a variant conformation consistent with a bend. Using an electrophoretic mobility shift assay, we demonstrated that Ner does not display significant affinity for a single 11-bp site. Furthermore, DNase I protection analysis and circular-permutation binding assays reveal that alterations in the length and sequence of the AT-rich spacer that separates the 11-bp inverted repeats significantly alter Ner-operator interactions, and demonstrate that the intrinsically bent ner operator is conformationally altered upon protein binding.