Escherichia coli K-12 outer membrane protein (OmpA) as a bacteriophage receptor: analysis of mutant genes expressing altered proteins

Abstract

The outer membrane protein OmpA of Escherichia coli K-12 serves as a receptor for a number of T-even-like phages. We have isolated a series of ompA mutants which are resistant to such phages but which still produce the OmpA protein. None of the mutants was able to either irreversibly or reversibly bind the phage with which they had been selected. Also, the OmpA protein is required for the action of colicins K and L and for the stabilization of mating aggregates in conjugation. Conjugal proficiency was unaltered in all cases. Various degrees of colicin resistance was found; however, the resistance pattern did not correlate with the phage resistance pattern. DNA sequence analyses revealed that, in the mutants, the 325-residue OmpA protein had suffered the following alterations: Gly-65-*Asp, Gly-65-Arg, Glu-68-Gly, Glu-68-Lys (two isolates), Gly-70-+Asp (four isolates), Gly-70-*Val, Ala-Asp-Thr-Lys-107-*Ala-Lys (caused by a 6-base-pair deletion), Val-110-Asp, and Gly-154-*Ser. These mutants exhibited a complex pattern of resistance-sensitivity to 14 different OmpA-specific phages, suggesting that they recognize different areas of the protein. In addition to the three clusters of mutational alterations around residues 68, 110, and 154, a site around residue 25 has been predicted to be involved in conjugation and in binding of a phage and a bacteriocin (R. Freudl, and S. T. Cole,

A molecular phylogeny of enteric bacteria and implications for a bacterial species concept

“…It could be the result of different genes experiencing different evolutionary pressures. For instance, gaps in the sequence alignment suggest that the ompA gene has experienced relatively strong selective pressure, presumably from bacteriophage such as K3 and Ox2, which have been shown to bind to the surface exposed loops of OmpA (Manning et al, 1976;Morona et al, 1984). Alternatively, the gene trees may differ because LGT has resulted in different phylogenetic histories for the different genes.…”

A molecular Swiss army knife: OmpA structure, function and expression

“…It was recognized soon after its discovery that it is a receptor for several bacteriophages, particularly phages K3, Ox2 and M1 (Schwarz et al, 1983;Morona et al, 1984;Riede et al, 1984;Montag et al, 1987). The p38 tail fibre protein of these viruses recognize OmpA through its surface-exposed loops 1, 2 and 3 (Morona et al, 1984(Morona et al, , 1985Koebnik, 1999). Recently, Power et al (2006) have identified two alleles of ompA, ompA1 and ompA2.…”

Conformation of Escherichia coli outer membrane protein OmpA produced in Bacillus subtilis: Influence of lipopolysaccharide

“…The bacteria were grown at 37°C with shaking (250 rpm), unless stated otherwise. The phage K3 [8,9] was propagated on the E. coli K-12 strain 701 [12], and treated with 0.5% Sarkosyl (N-lauroylsarcosine sodium salt, Sigma) which was subsequently replaced with 0.1% Triton X-100 by linear sucrose (5-20% w/w) gradient centrifugation (75 000 × g for 50 min at 10°C). The fraction with phage was diluted with antibiotic medium 3 as indicated.…”

“…The adjacent membrane-spanning/3-strands of OmpA form four surface loops around amino acid residues 25, 70, 110 and 154 [5]. These loops act as receptors for specific bacteriophage [8,9] and they are also needed to stabilize mating aggregates in F-mediated conjugation [8,10]. The /3-barrel structure applies only to the amino acid residues 1 to approx.…”

“…Four short peptide loops of OmpA are exposed on the surface of E. coli [5,9]. Two of these loops, the peptides around amino acid residues 70 and 110, are parts of the receptor for phage K3 [9]. The third loop around residue 154 is involved in conjugation [26].…”