Characterisation of holoenzyme lacking sigmaN regions I and II

The sigma-N (sigmaN) protein associates with bacterial core RNA polymerase to form a holoenzyme that is silent for transcription in the absence of enhancer-binding activator proteins. Here we show that the acidic Region II of sigmaN from Klebsiella pneumoniae is dispensable for polymerase isomerisation and trans-cription under conditions where the inhibited state of the holoenzyme is relieved by removal of sigmaN Region I sequences. Holoenzymes lacking Region I or Regions I+II were equally susceptible to the order of addition-dependent inhibition or stabilisation of DNA binding afforded by in trans Region I sequences. Region I+II-deleted [sigma] formed a holoenzyme with a DNA-binding activity more susceptible to inhibition by non-specific DNA than that lacking Region I. Region II sequences appear more closely associated with formation of a holoenzyme and [sigma] proficient in DNA binding than with changes in holoenzyme conformation needed for unmasking a single-strand DNA-binding activity used for open complex for-mation. Region II may therefore function to optimise DNA interactions for an efficient sigma cycle.