Immunochemical structure of the OmpD porin from Salmonella typhimurium

Abstract

The OmpD porin was isolated and purified from Salmonella typhimurium strain SH 7454 (ompC::TnlO), digested with cyanogen bromide (CNBr) and the peptide fragments were separated by SDS-PAGE. N-terminal sequencing identified a total of 96 residues from four distinct peptides. The sequence showed that OmpD is homologous to NmpC (75% identity), Lc (75%) and OmpC (70 YO) from Bcherichia coli, and OmpC (68%) from S. fjvphimurium . The sequence was essentially identical to the translated sequence of an nmpC-like gene of 5. typhimurium, currently placed at 3 8 6 centisomes of the chromosome. Our results and other data suggest, however, that this gene is actually the ompD gene, which is more correctly placed in the 34 centisome region of the chromosome. The CNBr-generated peptides were also screened with 16 anti-S. typhimurium OmpD monoclonal antibodies by Western blotting. These results, in conjunction with the prediction of the OmpD folding pattern based on the known three-dimensional structure of E. coli OmpF, showed a close immunological relationship among 5. fyphimurium OmpD and E. coli NmpC and Lc, and a strong conservation of sequences within the transmembrane Ip strands of these porins and E. coli OmpC, PhoE and OmpF, and Salmonella typhi OmpC.

A chromosomal region surrounding the ompD porin gene marks a genetic difference between Salmonella typhi and the majority of Salmonella serovars

“…Because both genes are cotransducible with phage P22, they cannot be that far apart. Instead, our results confirm those of Singh et al (1996) that place three S. typhimurium genes, ompD, smvA and narU, at centisome 33n7 (GenBank accession no. D26057).…”

“…Standard conditions for amplification were 30 cycles at 94 mC for 1 min, 55 mC for 1 min and 72 mC for 2 min, followed by a final extension step at 72 mC for 5 min. Primers OMPD1 (5h-GAC AAA GAC AAA ACC CGT T-3h) and OMPD2 (5h-CGT CCA GCA GGT TGA TTT T-3h) were used to amplify a 740 bp internal fragment of the ompD gene (Singh et al, 1996). Primers SMVA1 (5h-CTT AAC CGC CCG CTA TGA T-3h) and SMVA2 (5h-GCT GAA CCA CAT CCC TAC C-3h) were designed from the reported smvA sequence (GenBank accession no.…”

“…To determine whether the ompD gene is present in S. typhi, we first attempted to amplify a 740 bp fragment internal to ompD using PCR, with ompD-specific oligonucleotide primers (Singh et al, 1996). With these primers, no product was obtained when DNA isolated Absence of ompD in S. typhi from S. typhi strain Ty2, or DNA isolated from each of 26 independent S. typhi clinical isolates, was used as template (see Fig.…”

Contribution of Gene Amplification to Evolution of Increased Antibiotic Resistance inSalmonella typhimurium

“…Strain DA13421 (CE lineage 6) (in which bla TEM-1 gene copy number dropped) carried a missense mutation in the cpxA gene, the sensor part of another two-component system that regulates expression of OmpF and OmpC (Batchelor et al 2005). Finally, strain DA13513 (CF lineage 8) (in which bla TEM-1 copy number decreased slightly) carried a frameshift mutation in the nmpC gene, encoding the porin OmpD (Nikaido and Vaara 1985;Singh et al 1996).…”

Characterization of a novel gene related to antibiotic susceptibility in Pseudomonas aeruginosa

“…8,9 Some low-MW compounds can penetrate the outer membrane via porins, but larger, potentially harmful agents, including many antibiotics, are excluded. 10 As with antimicrobials, extensive use has led to increasing quinolone resistance among bacterial pathogens. Resistance to fluoroquinolones by P. aeruginosa is often mediated by mutations in target enzymes or upregulation in efflux pump expression, coupled with intrinsically low outer membrane permeability.…”