Detection and characterization of the S. typhimurium HilA protein

Christine R Rodriguez¹^,2 , Lisa M Schechter¹^,3 and Catherine A Lee¹

¹Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, MA 02115 USA

²Present address : Science Center, SC4186, Harvard University, Cambridge, MA 02138 USA

³Present address : Department of Plant Pathology, Cornell University, Ithaca, NY 14853 USA

author email corresponding author email

BMC Microbiology 2002, 2:31doi:10.1186/1471-2180-2-31


Published:	23 October 2002

Abstract

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.

Conclusion

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.