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, Eur. J. Biochem, 134:497-502, 1983; G. Braun and S. T. Cole, Mol. Gen. Genet, in press). These four areas are regularly spaced, being about 40 residues apart from each other. A model is suggested in which the OmpA polypeptide repeatedly traverses the outer membrane in cross-beta structure, exposing the four areas to the outside.
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Selected References
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- Bachmann B. J. Linkage map of Escherichia coli K-12, edition 7. Microbiol Rev. 1983 Jun;47(2):180–230. doi: 10.1128/mr.47.2.180-230.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Beck E., Bremer E. Nucleotide sequence of the gene ompA coding the outer membrane protein II of Escherichia coli K-12. Nucleic Acids Res. 1980 Jul 11;8(13):3011–3027. doi: 10.1093/nar/8.13.3011. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Benton W. D., Davis R. W. Screening lambdagt recombinant clones by hybridization to single plaques in situ. Science. 1977 Apr 8;196(4286):180–182. doi: 10.1126/science.322279. [DOI] [PubMed] [Google Scholar]
- Braun G., Cole S. T. Molecular characterization of the gene coding for major outer membrane protein OmpA from Enterobacter aerogenes. Eur J Biochem. 1983 Dec 15;137(3):495–500. doi: 10.1111/j.1432-1033.1983.tb07853.x. [DOI] [PubMed] [Google Scholar]
- Braun G., Cole S. T. The nucleotide sequence coding for major outer membrane protein OmpA of Shigella dysenteriae. Nucleic Acids Res. 1982 Apr 10;10(7):2367–2378. doi: 10.1093/nar/10.7.2367. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Braun V. Covalent lipoprotein from the outer membrane of Escherichia coli. Biochim Biophys Acta. 1975 Oct 31;415(3):335–377. doi: 10.1016/0304-4157(75)90013-1. [DOI] [PubMed] [Google Scholar]
- Bremer E., Beck E., Hindennach I., Sonntag I., Henning U. Cloned structural gene (ompA) for an integral outer membrane protein of Escherichia coli K-12: localization on hybrid plasmid pTU100 and expression of a fragment of the gene. Mol Gen Genet. 1980;179(1):13–20. doi: 10.1007/BF00268440. [DOI] [PubMed] [Google Scholar]
- Bremer E., Cole S. T., Hindennach I., Henning U., Beck E., Kurz C., Schaller H. Export of a protein into the outer membrane of Escherichia coli K12. Stable incorporation of the OmpA protein requires less than 193 amino-terminal amino-acid residues. Eur J Biochem. 1982 Feb;122(1):223–231. doi: 10.1111/j.1432-1033.1982.tb05870.x. [DOI] [PubMed] [Google Scholar]
- Chen R., Schmidmayr W., Krämer C., Chen-Schmeisser U., Henning U. Primary structure of major outer membrane protein II (ompA protein) of Escherichia coli K-12. Proc Natl Acad Sci U S A. 1980 Aug;77(8):4592–4596. doi: 10.1073/pnas.77.8.4592. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Clément J. M., Lepouce E., Marchal C., Hofnung M. Genetic study of a membrane protein: DNA sequence alterations due to 17 lamB point mutations affecting adsorption of phage lambda. EMBO J. 1983;2(1):77–80. doi: 10.1002/j.1460-2075.1983.tb01384.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cole S. T., Chen-Schmeisser U., Hindennach I., Henning U. Apparent bacteriophage-binding region of an Escherichia coli K-12 outer membrane protein. J Bacteriol. 1983 Feb;153(2):581–587. doi: 10.1128/jb.153.2.581-587.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cole S. T., Guest J. R. Genetic and physical characterization of lambda transducing phages (lambda frdA) containing the fumarate reductase gene of Escherichia coli K12. Mol Gen Genet. 1980;178(2):409–418. doi: 10.1007/BF00270492. [DOI] [PubMed] [Google Scholar]
- Cole S. T., Sonntag I., Henning U. Cloning and expression in Escherichia coli K-12 of the genes for major outer membrane protein OmpA from Shigella dysenteriae, Enterobacter aerogenes, and Serratia marcescens. J Bacteriol. 1982 Jan;149(1):145–150. doi: 10.1128/jb.149.1.145-150.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Datta D. B., Arden B., Henning U. Major proteins of the Escherichia coli outer cell envelope membrane as bacteriophage receptors. J Bacteriol. 1977 Sep;131(3):821–829. doi: 10.1128/jb.131.3.821-829.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Davies J. K., Reeves P. Genetics of resistance to colicins in Escherichia coli K-12: cross-resistance among colicins of group A. J Bacteriol. 1975 Jul;123(1):102–117. doi: 10.1128/jb.123.1.102-117.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dente L., Cesareni G., Cortese R. pEMBL: a new family of single stranded plasmids. Nucleic Acids Res. 1983 Mar 25;11(6):1645–1655. doi: 10.1093/nar/11.6.1645. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Endermann R., Krämer C., Henning U. Major outer membrane proteins of Escherichia coli K-12: evidence for protein II being a transmembrane protein. FEBS Lett. 1978 Feb 1;86(1):21–24. doi: 10.1016/0014-5793(78)80089-1. [DOI] [PubMed] [Google Scholar]
- Foulds J. Purification and partial characterization of a bacteriocin from Serratia marcescens. J Bacteriol. 1972 Jun;110(3):1001–1009. doi: 10.1128/jb.110.3.1001-1009.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Freudl R., Cole S. T. Cloning and molecular characterization of the ompA gene from Salmonella typhimurium. Eur J Biochem. 1983 Aug 15;134(3):497–502. doi: 10.1111/j.1432-1033.1983.tb07594.x. [DOI] [PubMed] [Google Scholar]
- Garavito R. M., Jenkins J. A., Neuhaus J. M., Pugsley A. P., Rosenbusch J. P. Structural investigations of outer membrane proteins from Escherichia coli. Ann Microbiol (Paris) 1982 Jan;133A(1):37–41. [PubMed] [Google Scholar]
- Hancock R. E., Reeves P. Bacteriophage resistance in Escherichia coli K-12: general pattern of resistance. J Bacteriol. 1975 Mar;121(3):983–993. doi: 10.1128/jb.121.3.983-993.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hancock R. E., Reeves P. Lipopolysaccharide-deficient, bacteriophage-resistant mutants of Escherichia coli K-12. J Bacteriol. 1976 Jul;127(1):98–108. doi: 10.1128/jb.127.1.98-108.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Havekes L. M., Hoekstra W. P. Characterization of an Escherichia coli K-12 F-Con-mutant. J Bacteriol. 1976 May;126(2):593–600. doi: 10.1128/jb.126.2.593-600.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Havekes L. M., Lugtenberg B. J., Hoekstra W. P. Conjugation deficient E. coli K12 F- mutants with heptose-less lipopolysaccharide. Mol Gen Genet. 1976 Jul 5;146(1):43–50. doi: 10.1007/BF00267981. [DOI] [PubMed] [Google Scholar]
- Heffron F., Bedinger P., Champoux J. J., Falkow S. Deletions affecting the transposition of an antibiotic resistance gene. Proc Natl Acad Sci U S A. 1977 Feb;74(2):702–706. doi: 10.1073/pnas.74.2.702. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Henning U., Hindennach I., Haller I. The major proteins of the Escherichia coli outer cell envelope membrane: evidence for the structural gene of protein II. FEBS Lett. 1976 Jan 1;61(1):46–48. doi: 10.1016/0014-5793(76)80168-8. [DOI] [PubMed] [Google Scholar]
- Henning U., Jann K. Two-component nature of bacteriophage T4 receptor activity in Escherichia coli K-12. J Bacteriol. 1979 Jan;137(1):664–666. doi: 10.1128/jb.137.1.664-666.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Henning U., Royer H. D., Teather R. M., Hindennach I., Hollenberg C. P. Cloning of the structural gene (ompA) for an integral outer membrane protein of Escherichia coli K-12. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4360–4364. doi: 10.1073/pnas.76.9.4360. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Henning U., Sonntag I., Hindennach I. Mutants (ompA) affecting a major outer membrane protein of Escherichia coli K12. Eur J Biochem. 1978 Dec;92(2):491–498. doi: 10.1111/j.1432-1033.1978.tb12771.x. [DOI] [PubMed] [Google Scholar]
- Hohn B. In vitro packaging of lambda and cosmid DNA. Methods Enzymol. 1979;68:299–309. doi: 10.1016/0076-6879(79)68021-7. [DOI] [PubMed] [Google Scholar]
- Humphreys G. O., Willshaw G. A., Smith H. R., Anderson E. S. Mutagenesis of plasmid DNA with hydroxylamine: isolation of mutants of multi-copy plasmids. Mol Gen Genet. 1976 Apr 23;145(1):101–108. doi: 10.1007/BF00331564. [DOI] [PubMed] [Google Scholar]
- Ish-Horowicz D., Burke J. F. Rapid and efficient cosmid cloning. Nucleic Acids Res. 1981 Jul 10;9(13):2989–2998. doi: 10.1093/nar/9.13.2989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kaiser K., Murray N. E. Physical characterisation of the "Rac prophage" in E. coli K12. Mol Gen Genet. 1979 Sep;175(2):159–174. doi: 10.1007/BF00425532. [DOI] [PubMed] [Google Scholar]
- Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
- Lugtenberg B., Peters R., Bernheimer H., Berendsen W. Influence of cultural conditions and mutations on the composition of the outer membrane proteins of Escherichia coli. Mol Gen Genet. 1976 Sep 23;147(3):251–262. doi: 10.1007/BF00582876. [DOI] [PubMed] [Google Scholar]
- Lugtenberg B., Van Alphen L. Molecular architecture and functioning of the outer membrane of Escherichia coli and other gram-negative bacteria. Biochim Biophys Acta. 1983 Mar 21;737(1):51–115. doi: 10.1016/0304-4157(83)90014-x. [DOI] [PubMed] [Google Scholar]
- Manning P. A., Puspurs A., Reeves P. Outer membrane of Escherichia coli K-12: isolation of mutants with altered protein 3A by using host range mutants of bacteriophage K3. J Bacteriol. 1976 Sep;127(3):1080–1084. doi: 10.1128/jb.127.3.1080-1084.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Manoil C. A genetic approach to defining the sites of interaction of a membrane protein with different external agents. J Mol Biol. 1983 Sep 15;169(2):507–519. doi: 10.1016/s0022-2836(83)80063-1. [DOI] [PubMed] [Google Scholar]
- Manoil C., Rosenbusch J. P. Conjugation-deficient mutants of Escherichia coli distinguish classes of functions of the outer membrane OmpA protein. Mol Gen Genet. 1982;187(1):148–156. doi: 10.1007/BF00384398. [DOI] [PubMed] [Google Scholar]
- Messing J., Crea R., Seeburg P. H. A system for shotgun DNA sequencing. Nucleic Acids Res. 1981 Jan 24;9(2):309–321. doi: 10.1093/nar/9.2.309. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Messing J., Vieira J. A new pair of M13 vectors for selecting either DNA strand of double-digest restriction fragments. Gene. 1982 Oct;19(3):269–276. doi: 10.1016/0378-1119(82)90016-6. [DOI] [PubMed] [Google Scholar]
- Morona R., Reeves P. Molecular cloning of the tolC locus of Escherichia coli K-12 with the use of transposon Tn10. Mol Gen Genet. 1981;184(3):430–433. doi: 10.1007/BF00352517. [DOI] [PubMed] [Google Scholar]
- Movva N. R., Nakamura K., Inouye M. Gene structure of the OmpA protein, a major surface protein of Escherichia coli required for cell-cell interaction. J Mol Biol. 1980 Nov 5;143(3):317–328. doi: 10.1016/0022-2836(80)90193-x. [DOI] [PubMed] [Google Scholar]
- Nakamura K., Mizushima S. Effects of heating in dodecyl sulfate solution on the conformation and electrophoretic mobility of isolated major outer membrane proteins from Escherichia coli K-12. J Biochem. 1976 Dec;80(6):1411–1422. doi: 10.1093/oxfordjournals.jbchem.a131414. [DOI] [PubMed] [Google Scholar]
- Nakamura K., Pirtle R. M., Inouye M. Homology of the gene coding for outer membrane lipoprotein within various Gram-negative bacteria. J Bacteriol. 1979 Jan;137(1):595–604. doi: 10.1128/jb.137.1.595-604.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Pierschbacher M. D., Ruoslahti E. Cell attachment activity of fibronectin can be duplicated by small synthetic fragments of the molecule. Nature. 1984 May 3;309(5963):30–33. doi: 10.1038/309030a0. [DOI] [PubMed] [Google Scholar]
- Puspurs A., Medon P., Corless C., Hackett J., Reeves P. A class of ompA mutants of Escherichia coli K12 affected in the interaction of ompA protein and the core region of lipopolysaccharide. Mol Gen Genet. 1983;189(1):162–165. doi: 10.1007/BF00326070. [DOI] [PubMed] [Google Scholar]
- Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
- Sanger F., Coulson A. R., Barrell B. G., Smith A. J., Roe B. A. Cloning in single-stranded bacteriophage as an aid to rapid DNA sequencing. J Mol Biol. 1980 Oct 25;143(2):161–178. doi: 10.1016/0022-2836(80)90196-5. [DOI] [PubMed] [Google Scholar]
- Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sarma V., Reeves P. Genetic locus (ompB) affecting a major outer-membrane protein in Escherichia coli K-12. J Bacteriol. 1977 Oct;132(1):23–27. doi: 10.1128/jb.132.1.23-27.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Schindler M., Osborn M. J. Interaction of divalent cations and polymyxin B with lipopolysaccharide. Biochemistry. 1979 Oct 2;18(20):4425–4430. doi: 10.1021/bi00587a024. [DOI] [PubMed] [Google Scholar]
- Schwarz H., Riede I., Sonntag I., Henning U. Degrees of relatedness of T-even type E. coli phages using different or the same receptors and topology of serologically cross-reacting sites. EMBO J. 1983;2(3):375–380. doi: 10.1002/j.1460-2075.1983.tb01433.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Schweizer M., Henning U. Action of a major outer cell envelope membrane protein in conjugation of Escherichia coli K-12. J Bacteriol. 1977 Mar;129(3):1651–1652. doi: 10.1128/jb.129.3.1651-1652.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Schweizer M., Hindennach I., Garten W., Henning U. Major proteins of the Escherichia coli outer cell envelope membrane. Interaction of protein II with lipopolysaccharide. Eur J Biochem. 1978 Jan 2;82(1):211–217. doi: 10.1111/j.1432-1033.1978.tb12013.x. [DOI] [PubMed] [Google Scholar]
- Skurray R. A., Hancock R. E., Reeves P. Con--mutants: class of mutants in Escherichia coli K-12 lacking a major cell wall protein and defective in conjugation and adsorption of a bacteriophage. J Bacteriol. 1974 Sep;119(3):726–735. doi: 10.1128/jb.119.3.726-735.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Van Alphen L., Havekes L., Lugtenberg B. Major outer membrane protein d of Escherichia coli K12. Purification and in vitro activity of bacteriophages k3 and f-pilus mediated conjugation. FEBS Lett. 1977 Mar 15;75(1):285–290. doi: 10.1016/0014-5793(77)80104-x. [DOI] [PubMed] [Google Scholar]
- Wahl G. M., Stern M., Stark G. R. Efficient transfer of large DNA fragments from agarose gels to diazobenzyloxymethyl-paper and rapid hybridization by using dextran sulfate. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3683–3687. doi: 10.1073/pnas.76.8.3683. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wilson G. G., Young K. Y., Edlin G. J., Konigsberg W. High-frequency generalised transduction by bacteriophage T4. Nature. 1979 Jul 5;280(5717):80–82. doi: 10.1038/280080a0. [DOI] [PubMed] [Google Scholar]
- Wilson J. H., Luftig R. B., Wood W. B. Interaction of bacteriophage T4 tail fiber components with a lipopolysaccharide fraction from Escherichia coli. J Mol Biol. 1970 Jul 28;51(2):423–434. doi: 10.1016/0022-2836(70)90152-x. [DOI] [PubMed] [Google Scholar]
- Yamamoto K. R., Alberts B. M., Benzinger R., Lawhorne L., Treiber G. Rapid bacteriophage sedimentation in the presence of polyethylene glycol and its application to large-scale virus purification. Virology. 1970 Mar;40(3):734–744. doi: 10.1016/0042-6822(70)90218-7. [DOI] [PubMed] [Google Scholar]
- Yu F., Mizushima S. Roles of lipopolysaccharide and outer membrane protein OmpC of Escherichia coli K-12 in the receptor function for bacteriophage T4. J Bacteriol. 1982 Aug;151(2):718–722. doi: 10.1128/jb.151.2.718-722.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]