Five families of outer membrane porins that function in protein secretion in Gram-negative bacteria are currently recognized. In this report, these five porin families are analyzed from structural and phylogenetic standpoints. They are the fimbrial usher protein (FUP), outer membrane factor (OMF), autotransporter (AT), two-partner secretion (TPS) and outer membrane secretin (Secretin) families. All members of these families in the current databases were identified, and all full-length homologues were multiply aligned for structural and phylogenetic analyses. The organismal distribution of homologues in each family proved to be unique with some families being restricted to proteobacteria and others being widespread in other bacterial kingdoms as well as eukaryotes. The compositions of and size differences between subfamilies provide evidence for specific orthologous relationships, which agree with available functional information and intra-subfamily phylogeny. The results reveal that horizontal transfer of genes encoding these proteins between phylogenetically distant organisms has been exceptionally rare although transfer within select bacterial kingdoms may have occurred. The resultant in silico analyses are correlated with available experimental evidence to formulate models relevant to the structures and evolutionary origins of these proteins.

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In Xanthomonas campestris, the causative agent of black rot in crucifers, the endoglucanase level is greatly decreased in the mutant deficient in Clp, a homologue of cyclic AMP receptor protein (CRP). It is established that Clp has the same DNA binding specificity as CRP at positions 5, 6, and 7 (GTG motif) of the DNA half site. In this study, the engA transcription initiation site was determined by the 5 0 RACE method, and two consensus Clp-binding sites, site I and site II centered at À69.5 and À42.5, respectively, were located. Transcriptional fusion assays indicated that Clp greatly activates engA transcription. Site-directed mutagenesis indicated that position 5 of GTG motif in site II is essential for both DNA-protein complex formation in electrophoretic mobility shift assays and engA transcription in vivo. In addition, mutation at position 5 of site I drastically reduces the promoter activity, indicating that binding of Clp to site I exerts a synergistic effect on the transcription activation by site II. engA appears to be the first X. campestris gene known to be activated by Clp via a direct binding to the promoter.

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Xanthomonas campestris pv. campestris possesses a low level of l-galactosidase and therefore is not able to grow and produce significant amounts of xanthan gum in a medium containing lactose as the sole carbon source. In this study, a ,I-galactosidase expression plasmid was constructed by ligating an X. campestris phage 4LO promoter with pKMOO5, a ColEl replicon containing Escherichia coli lacZY genes Rnd the lpp ribosome-binding site. It was then inserted into an IncPl broad-host-range plasmid, pLT, and subsequently transferred by conjugation to X. campestris 17, where it was stably maintained. The lacZ gene under the control of the phage promoter was expressed at a high level, enabling the cells to grow in a medium containing lactose. Production of xanthan gum in lactose or diluted whey by the engineered strain was evaluated, and it was found to produce as much xanthan gum in these substrates as the cells did in a medium containing glucose.