Cloned structural gene (ompA) for an integral outer membrane protein of Escherichia coli K-12 (original) (raw)
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Journal of Biomedical Science, 1998
Phage AR 1, previously known to infect Escherichia coli O 157:H7 with high specificity, was further characterized for its genetic properties. The phage DNA sequences including capsid genes and a putative a-glucosyltransferase gene (a-gt) have been deduced. These sequences are conservative but not identical to those of T4 phage. However, a nonessential gene, SegD, organized within the capsid gene cluster of T4 is missing in the corresponding region of AR1 genome, and this characteristic has not been observed among T-even related phages. The difference between AR1 and T4 was further exemplified by their distinct host ranges. Strains ofE. coli O 157:H7 collected from different sources were permissive to AR1 but resistant to T4 that normally infects K-12 strains of E. coli through contact with the outer membrane protein OmpC. Thus, the O157:H7 strains must have a varied OmpC. Indeed, the OmpC sequence of O157:H7 strains was proved to differ from that of K-12 strains by a total of 15 amino acid substitutions and two gaps (a five-residue deletion and a four-residue insertion). The OmpC molecules are relatively conserved across the gram-negative bacteria, and this is the first time OmpC divergence has been found within the same E. coli species. Since OmpC is located in the outer membrane and its expression is regulated by environmental conditions, alteration of the structure in pathogenic O 157:H7 strains may have biological significance. im oo,ol,llo m,*oot*allm
Nucleotide sequence of the gene ompA coding the outer membrane protein II * of Escherichia coli K-12
Nucleic Acids Research, 1980
A nucleotide sequence of 2271 basepairs has been determined from cloned E. ooli DNA which contains onpA. Within that sequence, starting at nucleotide 1037, an open translational reading frame encodes a protein of 367 amino acids which starting with amino acid 22 agrees with the primary structure of protein II*. The preceeding 21 amino acids constitute a typical signal sequence. There is a non-translated region of 360 nucleotides in front of the translational start. The insertion point of an IS1 element 110 nucleotides upstream from the start codon and an amber codon at the position of amino acid residue 28 have been localized in the DNA from two ompA mutants.
New pore protein produced in cells lysogenic for Escherichia coli phage HK253hrk
European Journal of Biochemistry, 1987
Outer membrane pore protein OmpC was identified as the receptor for the temperate Escherichia coli phage HK253hrk. The part of OmpC protein recognized by the phage was identified by using hybrid proteins in which parts of OmpC protein are replaced by the corresponding parts of the related PhoE protein. In contrast to other OmpC-specific phages, HK253hrk recognizes a part of OmpC within the C-terminal50 amino acids of the protein. E. coli strains lysogenic for HK253hrk produce reduced amounts of OmpC protein, and produce a new pore protein instead. Expression of this new protein was temperature-dependent, i. e. low at 30°C. The functioning of this new pore protein was characterized both in vivo by studying the uptake of P-lactam antibodies and in vitro after reconstitution of the protein in black lipid films. Its effective pore size was larger than that of the OmpF pores of E. coli B. The new porin appears to be cation-selective. A comparison with the selectivity of the known OmpC and OmpF pores of E. coli showed that the new pore has a higher selectivity than OmpF but is less selective than OmpC. The new pore protein appears to function in E. coli K12 lysogens as the receptor for the phages HK187, HK189 and HK332.
Molecular Microbiology, 1991
The kil-kor regulon of promiscuous plasmid RK2 includes the replication initiator gene trfA and several potentially host-lethal kif loci {kilA, kilB, ki/C, kilE), whose functions may be involved in plasmid maintenance or broad host range. The kilA locus consists of a single operon of three genes {klaA, ktaB, klaC), each of which is lethal when expressed from the klaA promoter in the absence of repressors encoded by korA and korB. In this study, we examined the effects of the unregulated klaA gene on the host cell. Bacteriophage X was used to construct a transducing phage {KpklaA-A) that allows efficient introduction of the klaA gene into Escherichia coli. Ceiis tacking korA and korB (to aliow uncontrolled expression of klaA) and expressing X repressor (to prevent phage lytic growth) are killed by XpkfaA-i. Cell death is dependent on the klaA structural gene, independent of the SOS system of the host, and is prevented by the presence of korA and korB. XpklaA-^ was used to synchronousiy infect ceils lacking korA and korB to determine the effects of klaA on the ceiis over time. The earliest effects, visible at two hours post-infection, are inhibition of growth of the culture, formation of eiongated cells, and striking changes in the appearance of the outer membrane. After four to five hours, the viability of the culture declined sharply and macromolecular synthesis ceased. The distinct ciass of early events is consistent with the hypothesis that the KlaA poiypeptide interacts with a specific target in the host cell.
Methods in Molecular Biology, 2009
Recent studies have established that the most abundant life form, that of phages, has had major influence on the biosphere, bacterial evolution, bacterial genome, and lateral gene transmission. Importantly the phages have served and continue to serve as valuable model systems. Such studies have led to a renewed interest and activity in the study of phages and their genomes. In order to determine the details of the involvement of phages in these important processes and activities, it is critical to assign specific functions to the phage gene products. The initial functional and gene assignments can be made by general mutagenesis of the phage genomes and of these specific gene products. A very informative mutagenic protocol that has found renewed interest is that using hydroxylamine. This mutagenic protocol has been used to obtain gene mutations involved in the lysogenic cycle of the Salmonella enterica serovar Anatum var. 15+ phage ε 34 (hereafter phage ε 34 ) and to isolate conditional lethal mutants of phage ε 34 . A similar protocol using plasmid is also described. A plate complementation method is presented to determine quickly the number of genes which are present in the population of mutations isolated from hydroxylamine mutagenesis.