EVIDENCE FOR SPECIFIC SECRETION RATHER THAN AUTOLYSIS IN THE RELEASE OF SOME HELICOBACTER-PYLORI PROTEINS

We investigated whether Helicobacter pylori cells actively secrete pro teins such as the urease subunits UreA and UreB and the GroES and GroE L homologs HspA and HspB or whether these proteins were present in the extracellular compartment as a consequence of autolysis. Using a subc ellular fractionation approach associated with quantitative Western bl ot analyses, we showed that the supernatant protein profiles were very different from those of the cell pellets, even for bacteria harvested in the late growth phase; this suggests that the release process is s elective. A typical cytoplasmic protein, a beta-galactosidase homolog, was found exclusively associated with the pellet of whole-cell extrac ts, and no traces were found in the supernatant. In contrast, UreA, Ur eB, HspA, and HspB were mostly found in the pellet but significant amo unts were also present in the supernatant. HspA and UreB were released into the supernatant at the same rate throughout the growth phase (3 %), whereas large portions of HspB and UreA were released during the s tationary phase (over 30 and 20 %, respectively) rather than during th e early growth phase (20 % and 6, respectively). The profiles of prote in obtained after water extraction of the bacteria with those of the p roteins naturally released within the liquid culture supernatants demo nstrated that water extraction led to the release of a large amount of protein due to artifactual lysis. Our data support the conclusion tha t a specific and selective mechanism(s) is involved in the secretion o f some H. pylori antigens. A programmed autolysis process does not see m to make a major contribution.