ABSTRACT
In Azospirillum brasilense, an extra-cytoplasmic function σ
factor (RpoE10) shows the characteristic 119 amino acid long C-terminal
extension found in ECF41-type σ factors, which possesses three conserved
motifs (WLPEP, DGGGR, and NPDKV), one in the linker region between the
σ2 and σ4, and the other two in the
SnoaL_2 domain of the C-terminal extension. Here, we have described the
role of the two conserved motifs in the SnoaL_2 domain of RpoE10 in the
inhibition and activation of its activity, respectively. Truncation
of the distal part of the C-terminal sequence of the RpoE10 (including
NPDKV but excluding the DGGGR motif) results in its promoter’s
activation suggesting autoregulation. Further truncation of the
C-terminal sequence up to its proximal part, including NPDKV and DGGGR
motif, abolished promoter activation. Replacement of NPDKV motif with
NAAAV in RpoE10 increased its ability to activate its promoter, whereas
replacement of DGGGR motif led to reduced promoter activation. We have
explored the dynamic modulation of 𝝈2 – 𝝈4 domains and the relevant
molecular interactions mediated by the two conserved motifs of the
SnoaL2 domain using molecular dynamics simulation. The analysis enabled
us to explain that the NPDKV motif located distally in the C-terminus
negatively impacts transcriptional activation. In contrast, the DGGGR
motif found proximally of the C-terminal extension is required to
activate RpoE10.