The σ subunit is the key regulator of bacterial transcription. Proteolysis of Thermus aquaticus σA, which occurred in situ during crystallization, reveals three domains, σ2, σ3, and σ4, connected by flexible linkers. Crystal structures of each domain were determined, as well as of σ4 complexed with −35 element DNA. Exposed surfaces of each domain are important for RNA polymerase binding. Universally conserved residues important for −10 element recognition and melting lie on one face of σ2, while residues important for extended −10 recognition lie on σ3. Genetic studies correctly predicted that a helix-turn-helix motif in σ4 recognizes the −35 element but not the details of the protein-DNA interactions. Positive control mutants in σ4 cluster in two regions, positioned to interact with activators bound just upstream or downstream of the −35 element.
