Detection and characterization of the S. typhimurium HilA protein.
BACKGROUND
Virulence genes on Salmonella pathogenicity island 1 (SPI1) are coordinately regulated by HilA, a member of the OmpR/ToxR family of transcription factors. Although a great deal is known about the complex regulation of hilA gene expression, very little is known about the HilA protein.


RESULTS
In order to detect and localize the HilA protein in S. typhimurium, we raised polyclonal antiserum against purified His-tagged HilA. This allowed us to study the effect of environmental conditions on the production of HilA. We also used the antiserum to examine the fractionation properties and SDS-PAGE mobility of native HilA. Our results indicate that S. typhimurium initiates translation of HilA from the first AUG codon in the hilA open-reading frame (ORF), producing a soluble 553 amino acid (63 kDa) protein product.


CONCLUSIONS
Materials and methods are now available to study the environmental regulation of the HilA protein in S. typhimurium. Our results also indicate that future in vitro studies of the interaction between HilA and DNA should utilize soluble preparations of HilA. Previous analyses used preparations of HilA in which the protein fractionated with the membrane, greatly limiting the types of experiments that could be conducted.
Rodriguez
Christine R
CR
Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, MA 02115 USA. christinerodriguez@post.harvard.edu
Schechter
Lisa M
LM
Lee
Catherine A
CA
D000332:Aerobiosis; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002458:Cell Fractionation; D004591:Electrophoresis, Polyacrylamide Gel; D015964:Gene Expression Regulation, Bacterial; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012486:Salmonella typhimurium / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*
D001426:Bacterial Proteins; C102236:HilA protein, Salmonella; D011993:Recombinant Fusion Proteins; C419942:Spi1 protein, Salmonella; D015534:Trans-Activators