Cassette mutagenesis implicates a helix-turn-helix motif in promoter recognition by the novel RNA polymerase sigma factor sigma 54.
Cassette mutagenesis has been used to study the role of a helix-turn-helix (HTH) motif in the novel RNA polymerase sigma factor sigma 54 of Klebsiella pneumoniae. Of the four residues which are predicted to be solvent-exposed in the second helix, the first (Glu-378) tolerated all substitutions, and some mutations of this residue increased expression from sigma 54-dependent promoters. Certain substitutions in the third exposed residue (Ser-382) produced a promoter-specific phenotype and all substitutions in the fourth residue (Arg-383) inactivated the protein, identifying this residue as being likely to be involved in base-specific interactions with the promoter. In vivo footprinting indicated that the inactive HTH mutants of sigma 54 were defective in interaction with both the -24 and -12 regions of the glnAp2 promoter.
Coppard
J R
JR
AFRC Nitrogen Fixation Laboratory, University of Sussex, Brighton, UK.
Merrick
M J
MJ
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics / Q000502:physiology; D007711:Klebsiella pneumoniae / Q000235:genetics*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D011401:Promoter Regions, Genetic / Q000502:physiology*; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D012808:Sigma Factor / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D011993:Recombinant Fusion Proteins; D012808:Sigma Factor; D001616:beta-Galactosidase