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. 1980 Aug;77(8):4499–4503. doi: 10.1073/pnas.77.8.4499

On the process of cellular division in Escherichia coli: isolation and characterization of penicillin-binding proteins 1a, 1b, and 3.

T Tamura, H Suzuki, Y Nishimura, J Mizoguchi, Y Hirota
PMCID: PMC349871  PMID: 7001458

Abstract

Multiple mutants of Escherichia coli defective in penicillin-binding proteins (PBPs) were constructed, and into these strains Co1E1 plasmids carrying the genes for PBP-1a, -1b, or -3 were introduced. From these plasmid-carrying strains, PBP-1a and -1b were purified by ampicillin-Sepharose affinity chromatography and PBP-3 by cephalexin-Sepharose affinity chromatography. Improved purification was achieved by differential elution with NH2OH. Purified PBP-1b synthesized murein when added to the membrane fraction of a PBP-1b-defective mutant, which by itself failed to support murein synthesis in vitro. The PBP-1b preparation was able to synthesize murein from the lipid intermediate extracted with chloroform/methanol but was unable to utilize UDP-linked precursors for murein synthesis. Murein synthesis was inhibited by vancomysin, ristocetin, moenomycin, and enduracidin, but not by beta-lactam antibiotics. The synthesized murein was shown to contain crosslinked muropeptides. Their crosslinking was abolished by action of beta-lactam antibiotics. The PBP-1a and -3 preparations showed substantially no activity for murein synthesis in the same reaction system. None of the three PBPs showed D-alanine carboxypeptidase activity with UDP-N-acetylmuramoyl-pentapeptide as substrate or endopeptidase activity with bis(disaccharide-peptide) as substrate.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anderson J. S., Matsuhashi M., Haskin M. A., Strominger J. L. Biosythesis of the peptidoglycan of bacterial cell walls. II. Phospholipid carriers in the reaction sequence. J Biol Chem. 1967 Jul 10;242(13):3180–3190. [PubMed] [Google Scholar]
  2. Blumberg P. M., Strominger J. L. Interaction of penicillin with the bacterial cell: penicillin-binding proteins and penicillin-sensitive enzymes. Bacteriol Rev. 1974 Sep;38(3):291–335. doi: 10.1128/br.38.3.291-335.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Blumberg P. M., Strominger J. L. Isolation by covalent affinity chromatography of the penicillin-binding components from membranes of Bacillus subtilis. Proc Natl Acad Sci U S A. 1972 Dec;69(12):3751–3755. doi: 10.1073/pnas.69.12.3751. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Clarke L., Carbon J. A colony bank containing synthetic Col El hybrid plasmids representative of the entire E. coli genome. Cell. 1976 Sep;9(1):91–99. doi: 10.1016/0092-8674(76)90055-6. [DOI] [PubMed] [Google Scholar]
  5. Georgopapadakou N., Hammarström S., Strominger J. L. Isolation of the penicillin-binding peptide from D-alanine carboxypeptidase of Bacillus subtilis. Proc Natl Acad Sci U S A. 1977 Mar;74(3):1009–1012. doi: 10.1073/pnas.74.3.1009. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Hartmann R., Höltje J. V., Schwarz U. Targets of penicillin action in Escherichia coli. Nature. 1972 Feb 25;235(5339):426–429. doi: 10.1038/235426a0. [DOI] [PubMed] [Google Scholar]
  7. Izaki K., Matsuhashi M., Strominger J. L. Glycopeptide transpeptidase and D-alanine carboxypeptidase: penicillin-sensitive enzymatic reactions. Proc Natl Acad Sci U S A. 1966 Mar;55(3):656–663. doi: 10.1073/pnas.55.3.656. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Jackson E. N., Yanofsky C. Localization of two functions of the phosphoribosyl anthranilate transferase of Escherichia coli to distinct regions of the polypeptide chain. J Bacteriol. 1974 Feb;117(2):502–508. doi: 10.1128/jb.117.2.502-508.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Kirschner K., Bisswanger H. Multifunctional proteins. Annu Rev Biochem. 1976;45:143–166. doi: 10.1146/annurev.bi.45.070176.001043. [DOI] [PubMed] [Google Scholar]
  10. Lugtenberg E. J., v Schijndel-van Dam A., van Bellegem T. H. In vivo and in vitro action of new antibiotics interfering with the utilization of N-acetyl-glucosamine-N-acetyl-muramyl-pentapeptide. J Bacteriol. 1971 Oct;108(1):20–29. doi: 10.1128/jb.108.1.20-29.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Matsuhashi M., Maruyama I. N., Takagaki Y., Tamaki S., Nishimura Y., Hirota Y. Isolation of a mutant of Escherichia coli lacking penicillin-sensitive D-alanine carboxypeptidase IA. Proc Natl Acad Sci U S A. 1978 Jun;75(6):2631–2635. doi: 10.1073/pnas.75.6.2631. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Matsuhashi M., Takagaki Y., Maruyama I. N., Tamaki S., Nishimura Y., Suzuki H., Ogino U., Hirota Y. Mutants of Escherichia coli lacking in highly penicillin-sensitive D-alanine carboxypeptidase activity. Proc Natl Acad Sci U S A. 1977 Jul;74(7):2976–2979. doi: 10.1073/pnas.74.7.2976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Mirelman D., Yashouv-Gan Y., Schwarz U. Peptidoglycan biosynthesis in a thermosensitive division mutant of Escherichia coli. Biochemistry. 1976 May 4;15(9):1781–1790. doi: 10.1021/bi00654a001. [DOI] [PubMed] [Google Scholar]
  14. Nishimura Y., Takeda Y., Nishimura A., Suzuki H., Inouye M., Hirota Y. Synthetic ColE1 plasmids carrying genes for cell division in Escherichia coli. Plasmid. 1977 Nov;1(1):67–77. doi: 10.1016/0147-619x(77)90009-9. [DOI] [PubMed] [Google Scholar]
  15. Oka T. Mode of action of penicillins in vivo and in vitro in Bacillus megaterium. Antimicrob Agents Chemother. 1976 Oct;10(4):579–591. doi: 10.1128/aac.10.4.579. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. PRIMOSIGH J., PELZER H., MAASS D., WEIDEL W. Chemical characterization of mucopeptides released from the E. coli B cell wall by enzymic action. Biochim Biophys Acta. 1961 Jan 1;46:68–80. doi: 10.1016/0006-3002(61)90647-3. [DOI] [PubMed] [Google Scholar]
  17. Shepherd S. T., Chase H. A., Reynolds P. E. The separation and properties of two penicillin-binding proteins from Salmonella typhimurium. Eur J Biochem. 1977 Sep;78(2):521–523. doi: 10.1111/j.1432-1033.1977.tb11765.x. [DOI] [PubMed] [Google Scholar]
  18. Spratt B. G. Distinct penicillin binding proteins involved in the division, elongation, and shape of Escherichia coli K12. Proc Natl Acad Sci U S A. 1975 Aug;72(8):2999–3003. doi: 10.1073/pnas.72.8.2999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Spratt B. G., Pardee A. B. Penicillin-binding proteins and cell shape in E. coli. Nature. 1975 Apr 10;254(5500):516–517. doi: 10.1038/254516a0. [DOI] [PubMed] [Google Scholar]
  20. Spratt B. G. Properties of the penicillin-binding proteins of Escherichia coli K12,. Eur J Biochem. 1977 Jan;72(2):341–352. doi: 10.1111/j.1432-1033.1977.tb11258.x. [DOI] [PubMed] [Google Scholar]
  21. Strominger J. L. Penicillin-sensitive enzymatic reactions in bacterial cell wall synthesis. Harvey Lect. 1968 1969;64:179–213. [PubMed] [Google Scholar]
  22. Suginaka H., Blumberg P. M., Strominger J. L. Multiple penicillin-binding components in Bacillus subtilis, Bacillus cereus, Staphylococcus aureus, and Escherichia coli. J Biol Chem. 1972 Sep 10;247(17):5279–5288. [PubMed] [Google Scholar]
  23. Suzuki H., Nishimura Y., Hirota Y. On the process of cellular division in Escherichia coli: a series of mutants of E. coli altered in the penicillin-binding proteins. Proc Natl Acad Sci U S A. 1978 Feb;75(2):664–668. doi: 10.1073/pnas.75.2.664. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Suzuki H., van Heijenoort Y., Tamura T., Mizoguchi J., Hirota Y., van Heijenoort J. In vitro peptidoglycan polymerization catalysed by penicillin binding protein 1b of Escherichia coli K-12. FEBS Lett. 1980 Feb 11;110(2):245–249. doi: 10.1016/0014-5793(80)80083-4. [DOI] [PubMed] [Google Scholar]
  25. Tamaki S., Nakajima S., Matsuhashi M. Thermosensitive mutation in Escherichia coli simultaneously causing defects in penicillin-binding protein-1Bs and in enzyme activity for peptidoglycan synthesis in vitro. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5472–5476. doi: 10.1073/pnas.74.12.5472. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Tamura T., Imae Y., Strominger J. L. Purification to homogeneity and properties of two D-alanine carboxypeptidases I From Escherichia coli. J Biol Chem. 1976 Jan 25;251(2):414–423. [PubMed] [Google Scholar]
  27. Tipper D. J., Strominger J. L. Mechanism of action of penicillins: a proposal based on their structural similarity to acyl-D-alanyl-D-alanine. Proc Natl Acad Sci U S A. 1965 Oct;54(4):1133–1141. doi: 10.1073/pnas.54.4.1133. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Van Heijenoort Y., Derrien M., Van Heijenoort J. Polymerization by transglycosylation in the biosynthesis of the peptidoglycan of Escherichia coli K 12 and its inhibition by antibiotics. FEBS Lett. 1978 May 1;89(1):141–144. doi: 10.1016/0014-5793(78)80540-7. [DOI] [PubMed] [Google Scholar]
  29. Wise E. M., Jr, Park J. T. Penicillin: its basic site of action as an inhibitor of a peptide cross-linking reaction in cell wall mucopeptide synthesis. Proc Natl Acad Sci U S A. 1965 Jul;54(1):75–81. doi: 10.1073/pnas.54.1.75. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Yocum R. R., Waxman D. J., Rasmussen J. R., Strominger J. L. Mechanism of penicillin action: penicillin and substrate bind covalently to the same active site serine in two bacterial D-alanine carboxypeptidases. Proc Natl Acad Sci U S A. 1979 Jun;76(6):2730–2734. doi: 10.1073/pnas.76.6.2730. [DOI] [PMC free article] [PubMed] [Google Scholar]

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