Characterisation of holoenzyme lacking sigma(N) regions I and II

The sigma-N (sigma(N)) protein associates with bacterial core RNA polymeras e to form a holoenzyme that is silent for transcription in the absence of e nhancer-binding activator proteins. Here we show that the acidic Region II of sigma(N) from Klebsiella pneumoniae is dispensable for polymerase isomer isation and transcription under conditions where the inhibited state of the holoenzyme is relieved by removal of sigma(N) Region I sequences. Holoenzy mes lacking Region I or Regions I+II were equally susceptible to the order of addition-dependent inhibition or stabilisation of DNA binding afforded b y in trans Region I sequences. Region I+II-deleted sigma formed a holoenzym e with a DNA-binding activity more susceptible to inhibition by non-specifi c DNA than that lacking Region I. Region Ii sequences appear more closely a ssociated with formation of a holoenzyme and sigma proficient in DNA bindin g than with changes in holoenzyme conformation needed for unmasking a singl e-strand DNA-binding activity used for open complex formation. Region II ma y therefore function to optimise DNA interactions for an efficient sigma cy cle.