IMMUNOLOGICAL AND MOLECULAR CHARACTERIZATION OF HELICOBACTER-FELIS UREASE

Urease activity has recently been shown to be an important virulence d eterminant for Helicobacter pylori, allowing it to survive the low pH of the stomach during colonization. Experimental murine infection with Helicobacter felis is now being used as a model for H. pylori infecti on to study the effects of vaccines, antibiotics, and urease inhibitor s on colonization. However, little information comparing the ureases o f H. felis and H. pylori is available. Urease was partially purified f rom the cell surface of H. felis ATCC 49179 by A-5M agarose chromatogr aphy, resulting in an eightfoId increase in specific activity over tha t of crude urease. The apparent K-m for urea for the partially purifie d urease was 0.4 mM, and the enzyme was inhibited in a competitive man ner by flurofamide (50% inhibitory concentration = 0.12 mu M). Antiser um to whole cells of H. pylori recognized both H. pylori and H. felis urease B subunits. Antiserum raised against H. felis whole cells recog nized the large and small autologous urease subunits and the cpn60 hea t shock molecule in both H. felis and H. pylori. However, this antiser um showed only a weak reaction with the B subunit of H. pylori urease. Two oligomeric DNA sequences were used as probes to evaluate the rela tedness of H. felis and H. pylori urease gene sequences. One 30-mer fr om the ureA sequence, which had been shown previously to be specific f or H. pylori, failed to hybridize to H. felis genomic DNA. A probe to the putative coding sequence for the active site of the H. pylori ureB subunit hybridized at low intensity to a 2.8-kb fragment of BamHI-Hin dIII-digested H. felis DNA, suggesting that the sequences were homolog ous but not identical, a result confirmed from the recently published sequences of ureA and ureB from H. felis.