UTILIZATION OF ALKALINE-PHOSPHATASE FUSIONS TO IDENTIFY SECRETED PROTEINS, INCLUDING POTENTIAL EFFLUX PROTEINS AND VIRULENCE FACTORS FROM HELICOBACTER-PYLORI

The targeted genomic strategy of random fusions to a partial gene enco ding a signal sequence-deficient fragment of bacterial alkaline phosph atase was utilized to screen for secreted proteins in Helicobacter pyl ori. The rationale for targeting extracytoplasmic proteins was based o n the hypothesis that most virulence factors and vaccine candidates ar e secreted or exported proteins. In addition, extracytosolic proteins represent good potential targets for drug intervention since they are in general more accessible to drugs than are cytoplasmically localized proteins. The application of this strategy to H. pylori allowed the i dentification of putative virulence factors and novel targets for drug intervention including four putative antibiotic efflux genes. The str ategy used here is rapid and technically simple, relatively inexpensiv e, adaptable to a wide variety of microbes and genetic systems, and se lects for expressed and accessible proteins.