Characterisation of a transitionally occupied state of domain 1.1 of σA
factor of RNA polymerase from Bacillus subtilis
Abstract
σ factors are essential parts of bacterial RNA polymerase (RNAP)
as they allow to recognize promotor sequences and initiate
transcription. Domain 1.1 of vegetative σ factors occupies the
primary channel of RNAP and also prevents binding of the σ
factor to promoter DNA alone. Here, we show that domain 1.1 of
Bacillus subtilis σ A exists in two structurally distinct
variants in dynamic equilibrium. The major conformation at room
temperature is represented by a previously reported well-folded
structure solved by nuclear magnetic resonance (NMR), but 4% of the
protein molecules are present in a less thermodynamically favorable
state. We show that this population increases with temperature and may
represent as much as 20% at 43.5 ◦ C. We characterized the
minor state of the domain 1.1 using specialized methods of NMR. We found
that, in contrast to the major state, the detected minor state is
partially unfolded. Its propensity to form secondary structure elements
is especially decreased for the first and third α helices,
while the second α helix and β strand close to the
C-terminus are more stable. In summary, this study reveals
conformational dynamics of domain 1.1 and provides a basis for studies
of its interaction with RNAP and effects on transcription regulation.
