Characterization of omp200, a porin gene complex from Bacteroides fragilis:: omp121 and omp71, gene sequence, deduced amino acid sequences and predictions of porin structure

The high MW porin protein complex (Omp200. composed of Omp121 and Omp71) from Bacteroides fragilis ATCC 25285 was purified and tryptic peptide sequences were used to design degenerate oligonucleotide primers which A ere then used as a first step in amplification, identification and sequencing of the omp121 gene (GenBank Accession Number AF357210). Sequence analysis revealed an open reading frame of 3378 bases. The deduced amino acid sequence (which contained the experimentally determined peptide sequences) has 1125 or 1116 amino acids (depending on which start codon is used): the mature protein consists of 1096 amino acids, has a predicted MW of 121.4 and a theoretical pI of 6,32. It is preceded by a 29 or 18 amino acid signal peptide which includes a typical hydrophobic region near the N- terminus (VLVLVL), Hydropathy plots of the deduced amino acid sequence of B. fragilis Omp 121 display striking similarity A with those of Escherichia coli OmpC (a 16-stranded porin) and FepA (a 22-stranded ligand-gated transport protein). Three-dimensional modeling of B, fragilis Omp121 (based on 1D and 3D sequence profiles. coupled with secondary structure and solvation potential information) indicated that the closest homologues in terms in fold conservation were the E. coli 16-stranded porins (e.g. OsmA) and 22-stranded ligand gated transport proteins (e.g. FepA). The omp71 gene sequence was identified using the tryptic peptides to search the published Bacteroides genome data base. We found that omp71 is located immediately downstream of omp121 and confirmed this with PCR analysis. Omp71 has no known homologues but does share some characteristics with the Porphyromonas RagB antigen. (C) 2002 Elsevier Science B.V. All rights reserved.