Functional analysis of the Helicobacter pylori principal sigma subunit of RNA polymerase reveals that the spacer region is important for efficient transcription.
We have cloned the rpoD gene encoding the principal sigma (sigma) factor of Helicobacter pylori. The deduced amino acid sequence reveals a predicted polypeptide of 676 residues that has amino acid homology with the principal sigma factors of a number of divergent prokaryotes. We have designated this factor sigma80. Amino acid sequence analysis suggests that region 1.1 is missing in sigma80 and that a region with homology to a regulatory protein from Bacillus subtilis phage SPO1 is present. Genetic studies have indicated that sigma80 is not compatible with the transcriptional machinery of Escherichia coli. However, in vitro sigma80 could be assembled into the E. coli RNA polymerase and could bind to E. coli and H. pylori promoters, suggesting that the sigma80-containing RNA polymerase has the same stoichiometry as the native complex. By exchanging protein domains between E. coli and H. pylori sigma factors, we demonstrate that the sigma80 domain inhibiting transcription from E. coli promoters is confined within the non-conserved spacer region, implying that the spacer region of prokaryotic primary sigma factors plays an important role in the process of transcription. Consistent with its restricted niche and with the availability of a very restricted number of transcriptional regulators, H. pylori may have evolved a spacer region of the sigma factor to modulate total transcription and to quickly respond to microenvironmental changes.
Beier
D
D
Department of Molecular Biology, Chiron SpA, IRIS Research Institute, Siena, Italy.
Spohn
G
G
Rappuoli
R
R
Scarlato
V
V
D000595:Amino Acid Sequence; D000942:Antigens, Bacterial; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D016480:Helicobacter pylori / Q000201:enzymology / Q000235:genetics*; D020035:Holoenzymes / Q000378:metabolism; D015151:Immunoblotting; D007763:Lac Operon; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D012808:Sigma Factor / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D014158:Transcription, Genetic
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D020035:Holoenzymes; D011994:Recombinant Proteins; D012808:Sigma Factor; C080866:cagA protein, Helicobacter pylori; C081775:RNA polymerase sigma 70; D012321:DNA-Directed RNA Polymerases