Apparent bacteriophage-binding region of an Escherichia coli K-12 outer membrane protein

Abstract

The 325-residue OmpA protein is one of the major outer membrane proteins of Escherichia coli. It serves as the receptor for several T-even-like phages and is required for the action of certain colicins and for the stabilization of mating aggregates in conjugation. We have isolated two mutant alleles of the cloned ompA gene which produce a protein that no longer functions as a phage receptor. Bacteria possessing the mutant proteins were unable to bind the phages, either reversibly or irreversibly. However, both proteins still functioned in conjugation, and one of them conferred colicin L sensitivity. DNA sequence analysis showed that the phage-resistant, colicin-sensitive phenotype exhibited by one mutant was due to the amino acid substitution Gly --Arg at position 70. The second mutant,

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

“…The phage-resistant ompA mutants analyzed possess alterations at three areas of the OmpA protein: regions 70, 110, and 154. Most of the mutational alterations we found were located at region 70 (13 of 16, including the two mutants analyzed previously [13]). For this region, direct evidence that it is exposed at the cell surface has been obtained (13).…”

“…The plasmids PTU105 and pTU115 have been described before (13). pTU202 and PTU203 were obtained by hydroxylamine mutagenesis (35) of pTU201, which contains the complete ompA gene cloned into pBR325 (8), followed by transformation into strain UH201.3 and selection for resistance to phage Ox2.…”

“…The results are shown in Table 2. For comparison, the previously characterized mutants (ompA105 and ompA115 [13]) are also included.…”

Bacteriophage receptor area of outer membrane protein OmpA of Escherichia coli K-12

“…Reversible adsorption was tested by incubating a mixture of such cells consisting of a threefold excess of mutant over wild-type cells with phage. The phage neutralization rate was compared with that determined for wild-type cells alone (12,20). Strain RMT182, lacking detectable amounts of OmpA protein, was used as a negative control.…”

“…The results suggested that the protein is exposed at the surface of the cell at least around residues 25, 70, and 110 (numbers according to the E. coli sequence). In another, we have analyzed phage-resistant ompA mutants which produce an altered protein (12,20). The mutant proteins had alterations affecting three regions of the polypeptide, located around amino acid residues 70, 110, and 154.…”