Helicobacter pylori with separate beta- and beta'-subunits of RNA polymerase is viable and can colonize conventional mice.
The genes encoding the beta- and beta'-subunits of RNA polymerase (rpoB and rpoC respectively) are fused as one continuous open reading frame in Helicobacter pylori and in other members of this genus, but are separate in other bacterial taxonomic groups, including the closely related genus Campylobacter. To test whether this beta-beta' tethering is essential, we used polymerase chain reaction-based cloning to separate the rpoB and rpoC moieties of the H. pylori rpoB-rpoC fusion gene with a non-polar chloramphenicol resistance cassette containing a new translational start, and introduced this construct into H. pylori by electro-transformation. H. pylori containing these separated rpoB and rpoC genes in place of the native fusion gene produced non-tethered beta and beta' RNAP subunits, grew well in culture and colonized and proliferated well in conventional C57BL/6 mice. Thus, the extraordinary beta-beta' tethering is not essential for H. pylori viability and gastric colonization.
Raudonikiene
A
A
Department of Molecular Microbiology, Washington University School of Medicine, St Louis 63110, MO, USA.
Zakharova
N
N
Su
W W
WW
Jeong
J Y
JY
Bryden
L
L
Hoffman
P S
PS
Berg
D E
DE
Severinov
K
K
D000818:Animals; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D003001:Cloning, Molecular; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D018274:Electroporation; D005260:Female; D016480:Helicobacter pylori / Q000201:enzymology* / Q000472:pathogenicity*; D051379:Mice / Q000382:microbiology; D008810:Mice, Inbred C57BL; D008957:Models, Genetic; D016254:Mutagenesis, Insertional; D010940:Plant Proteins / Q000235:genetics; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins; D013270:Stomach / Q000382:microbiology; D013997:Time Factors; D014170:Transformation, Genetic
D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; C101305:RpoB protein, Oryza sativa; D015500:Chloramphenicol O-Acetyltransferase; C108570:beta' subunit of RNA polymerase; D012321:DNA-Directed RNA Polymerases