MOLECULAR CHARACTERIZATION OF THE MAJOR OUTER-MEMBRANE PROTEIN OPRF FROM PLANT ROOT-COLONIZING PSEUDOMONAS-FLUORESCENS

N-terminal sequence analysis of peptides generated by proteolytic trea tment of the Pseudomonas fluorescens OE 28.3 major outer-membrane prot ein OprF, embedded in outer membranes or present in whole cells, indic ated a surface-exposed location for the proline-rich region of the pro tein. This region is absent from the P. aeruginosa and P. syringae Opr Fs. Evidence was obtained for the presence of additional exposed but l ess accessible regions in the carboxy half of OprF. Four OprF-specific monoclonal antibodies were all directed to the C-terminal part of the protein but did not recognize a surface-exposed epitope as shown by f low cytometry. Our data support the model previously proposed for P. a eruginosa OprF in which the entire protein is embedded in the outer me mbrane, unlike the topology proposed for the major outer-membrane prot ein from Escherichia coli, OmpA, whose carboxy half resides in the per iplasmic space. For six other P. fluorescens strains producing OprF pr oteins with different isoelectric paints, the primary structure was de termined by sequence analysis of the PCR-amplified oprF genes. The pro line-rich domain represented the most conserved region of the differen t P. fluorescens OprFs. Based on the sequence of its oprF gene, it was shown that the mushroom pathogen P. tolaasii is quite closely related to P. fluorescens. Comparative sequence analysis further showed that the carboxy half of OprF contains a sequence motif that is well conser ved in the enterobacterial OmpA proteins but is also present in a numb er of other outer-membrane proteins, including peptidoglycan-associate d lipoproteins.