Two domains within σ ^N (σ ⁵⁴ ) cooperate for DNA binding

Analysis of the architecture of the transcription factor σN (σ54) and its domains by circular dichroism

“…Although the critical residues within the 425-477 deletion interval responsible for the markedly altered melting behaviour of the C-terminally truncated protein are not defined, the so-called RPON box, a conserved sequence of eight amino acids in the C-terminus of N proteins, is a candidate structural element in N (Cannon et al, 1995a;Merrick, 1993;Taylor et al, 1996). Since interactions between residues 329-477 and 180-306 are important for DNA-binding (Cannon et al, 1997) these interactions may be reflected in the altered melting transitions discussed above. Conformational adjustments and inherent dynamic changes are expected of N as it participates in the process required for activation of transcription, in particular protein-and DNA-binding events.…”

Amino-terminal sequences of sigma N (sigma 54) inhibit RNA polymerase isomerization

“…Proteins were prepared as described previously (Cannon et al 1995b(Cannon et al , 1997Casaz and Buck 1997 Buck, in prep.). A purified carboxy-terminally deleted form of activator, PspF⌬HTH (Jovanovic et al 1996) was used in activation assays.…”

DNA‐binding domain mutants of sigma‐N (σN, σ54) defective between closed and stable open promoter complex formation

“…The N -holoenzyme appears poised for melting since, in the closed complex: (i) sequences melted in the open complex are distorted; and (ii) nucleation of strand separation around the ¹12 promoter element occurs (Morris et al, 1994;Wang et al, 1997). Contacts to promoter DNA are made through C-terminal N sequences Cannon et al, 1995a; and at least five discrete sequences associated with DNA binding have been identified (Taylor et al, 1996;Guo and Gralla, 1997 DNA binding Cannon et al, 1997;Missailidis et al, 1997). Thus, several domains may create an active centre in N associated with DNA recognition.…”