AEROMONAS-SALMONICIDA POSSESSES 2 GENES ENCODING HOMOLOGS OF THE MAJOR OUTER-MEMBRANE PROTEIN, OMPA

Two homologs of the outer membrane protein OmpA were identified in Aer omonas salmonicida by sodium dodecyl sulfate-polyacrylamide gel electr ophoresis, immunoblotting, and amino-terminal sequence analyses, An A. salmonicida genomic DNA library was constructed by using lambda GEM-1 1 and recombinant phage carrying both genes (ompAI and ompAII) selecte d by immunoscreening. A 5.0-kb BamHI fragment containing the two genes in tandem was subcloned in pBluescript and used for further subclonin g and sequencing of the genes, The encoded proteins (M(r) = 33,564 and 32,536 for mature OmpAI and OmpAII, respectively) had only 64% identi ty with each other and otherwise had the highest level of homology to OmpA proteins from the members of the family Enterobacteriaceae. Based on the Escherichia coli OmpA model, an eight-stranded amphipathic bet a-barrel model for the membrane assembly of the N-terminal half of Omp AI and OmpAII was predicted. Most variation between the two proteins w as localized to the predicted surface loops and periplasmic turns, whi le the transmembrane strands and C-terminal domains were highly conser ved, Expression of ompAI and ompAII separately in E. coli indicated th at both genes could be independently transcribed from their own promot ers and that both gene products were assembled into the E. coli outer membrane, A survey of different Aeromonas spp. by PCR revealed that po ssession of two tandem ompA genes was widespread among this genus, Thi s is the first report of any bacterial species possessing two genes fo r homologs of this major outer membrane protein.