DNA Recognition by a σ54 Transcriptional Activator from Aquifex aeolicus

Transcription initiation by bacterial sigma(54)-polymerase requires the action of a transcriptional activator protein. Activators bind sequence-specifically upstream of the transcription initiation site via a DNA-binding domain (DBD). The structurally characterized DBDs from activators all belong to the Fis (factor for inversion stimulation) family of helix-turn-helix DNA-binding proteins. We report here structures of the free and DNA-bound forms of the DBD of NtrC4 (4DBD) from Aquifex aeolicus, a member of the NtrC family of sigma(54) activators. Two NtrC4-binding sites were identified upstream (-145 and -85 bp) from the start of the IpxC gene, which is responsible for the first committed step in lipid A biosynthesis. This is the first experimental evidence for sigma(54) regulation in IpxC expression. 4DBD was crystallized both without DNA and in complex with the - 145-binding site. The structures, together with biochemical data, indicate that NtrC4 binds to DNA in a manner that is similar to that of its close homolog, Fis. The greater sequence specificity for the binding of 4DBD relative to Fis seems to arise from a larger number of base-specific contacts contributing to affinity than for Fis. (C) 2014 Elsevier Ltd. All rights reserved.