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. 1982 Mar;149(3):880–888. doi: 10.1128/jb.149.3.880-888.1982

Kinetics of Biosynthesis of Iron-Regulated Membrane Proteins in Escherichia coli

Phillip E Klebba 1, Mark A McIntosh 1, J B Neilands 1
PMCID: PMC216474  PMID: 6174499

Abstract

Using biological iron chelators to control specifically iron availability to Escherichia coli K-12 in conjunction with radioactive pulse-labels, we examined the biosynthesis of six iron-regulated membrane proteins. Iron deprivation induced the synthesis of five proteins, which had molecular weights of 83,000 (83K), 81K (Fep), 78K (TonA), 74K (Cir), and 25K. The kinetics of induction were the same in entA and entA+ strains, but were affected by the initial iron availability in the media. Iron-poor cells induced rapidly (half-time, 10 min), whereas iron-rich cells began induction after a lag and showed a slower induction half-time (30 min). Within this general pattern of induction after iron deprivation, several different kinetic patterns were apparent. The 83K, 81K, and 74K proteins were coordinately controlled under all of the conditions examined. The 78K and 25K proteins were regulated differently. The synthesis of a previously unrecognized 90K inner membrane protein was inhibited by iron deprivation and stimulated by iron repletion. Both ferrichrome and ferric enterobactin completely repressed 81K and 74K synthesis when the siderophores were supplied at concentrations of 5 μM in vivo (half-time, 2.5 min). At concentrations less than 5 μM, however, both siderophores repressed synthesis only temporarily; the duration of repression was proportional to the amount of ferric siderophore added. The half-lives of the 81K and 74K mRNAs, as measured by rifampin treatment, were 1.2 and 1.6 min, respectively. The results of this study suggest that enteric bacteria are capable of instantaneously detecting and reacting to fluctuations in the extracellular iron concentration and that they store iron during periods of iron repletion for utilization during periods of iron stress. Neither iron storage nor iron regulation of envelope protein synthesis is dependent on the ability of the bacteria to form heme.

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Selected References

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  1. Ames G. F. Resolution of bacterial proteins by polyacrylamide gel electrophoresis on slabs. Membrane, soluble, and periplasmic fractions. J Biol Chem. 1974 Jan 25;249(2):634–644. [PubMed] [Google Scholar]
  2. Bachmann B. J., Low K. B. Linkage map of Escherichia coli K-12, edition 6. Microbiol Rev. 1980 Mar;44(1):1–56. doi: 10.1128/mr.44.1.1-56.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bartlett B. O., Scott Russell R., Jenkins W. Improved relationship between the deposition of strontium-90 and the contamination of milk in the United Kingdom. Nature. 1972 Jul 7;238(5358):46–48. doi: 10.1038/238046a0. [DOI] [PubMed] [Google Scholar]
  4. Bauminger E. R., Cohen S. G., Labenski de Kanter F., Levy A., Ofer S., Kessel M., Rottem S. Iron storage in Mycoplasma capricolum. J Bacteriol. 1980 Jan;141(1):378–381. doi: 10.1128/jb.141.1.378-381.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bonner W. M., Laskey R. A. A film detection method for tritium-labelled proteins and nucleic acids in polyacrylamide gels. Eur J Biochem. 1974 Jul 1;46(1):83–88. doi: 10.1111/j.1432-1033.1974.tb03599.x. [DOI] [PubMed] [Google Scholar]
  6. Boyd A., Holland I. B. Protein d, an iron-transport protein induced by filtration of cultures of Escherichia coli. FEBS Lett. 1977 Apr 1;76(1):20–24. doi: 10.1016/0014-5793(77)80112-9. [DOI] [PubMed] [Google Scholar]
  7. Braun V., Hancock R. E., Hantke K., Hartmann A. Functional organization of the outer membrane of escherichia coli: phage and colicin receptors as components of iron uptake systems. J Supramol Struct. 1976;5(1):37–58. doi: 10.1002/jss.400050105. [DOI] [PubMed] [Google Scholar]
  8. Braun V., Schaller K., Wolff H. A common receptor protein for phage T5 and colicin M in the outer membrane of Escherichia coli B. Biochim Biophys Acta. 1973 Sep 27;323(1):87–97. doi: 10.1016/0005-2736(73)90433-1. [DOI] [PubMed] [Google Scholar]
  9. Cardelli J., Konisky J. Isolation and characterization of an Escherichia coli mutant tolerant to colicins Ia and Ib. J Bacteriol. 1974 Aug;119(2):379–385. doi: 10.1128/jb.119.2.379-385.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Cox G. B., Gibson F., Luke R. K., Newton N. A., O'Brien I. G., Rosenberg H. Mutations affecting iron transport in Escherichia coli. J Bacteriol. 1970 Oct;104(1):219–226. doi: 10.1128/jb.104.1.219-226.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Guterman S. K. Inhibition of colicin B by enterochelin. Biochem Biophys Res Commun. 1971 Sep;44(5):1149–1155. doi: 10.1016/s0006-291x(71)80206-1. [DOI] [PubMed] [Google Scholar]
  12. Hancock R. E., Hantke K., Braun V. Iron transport of Escherichia coli K-12: involvement of the colicin B receptor and of a citrate-inducible protein. J Bacteriol. 1976 Sep;127(3):1370–1375. doi: 10.1128/jb.127.3.1370-1375.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Harrison P. M. Iron storage in bacteria. Nature. 1979 May 3;279(5708):15–16. doi: 10.1038/279015a0. [DOI] [PubMed] [Google Scholar]
  14. Hirashima A., Childs G., Inouye M. Differential inhibitory effects of antibiotics on the biosynthesis of envelope proteins of Escherichia coli. J Mol Biol. 1973 Sep 15;79(2):373–389. doi: 10.1016/0022-2836(73)90012-0. [DOI] [PubMed] [Google Scholar]
  15. Hollifield W. C., Jr, Neilands J. B. Ferric enterobactin transport system in Escherichia coli K-12. Extraction, assay, and specificity of the outer membrane receptor. Biochemistry. 1978 May 16;17(10):1922–1928. doi: 10.1021/bi00603a019. [DOI] [PubMed] [Google Scholar]
  16. Jacquet M., Kepes A. Initiation, elongation and inactivation of lac messenger RNA in Escherichia coli studied studied by measurement of its beta-galactosidase synthesizing capacity in vivo. J Mol Biol. 1971 Sep 28;60(3):453–472. doi: 10.1016/0022-2836(71)90181-1. [DOI] [PubMed] [Google Scholar]
  17. KEPES A. KINETICS OF INDUCED ENZYME SYNTHESIS. DETERMINATION OF THE MEAN LIFE OF GALACTOSIDASE-SPECIFIC MESSENGER RNA. Biochim Biophys Acta. 1963 Oct 15;76:293–309. [PubMed] [Google Scholar]
  18. Laskey R. A., Mills A. D. Quantitative film detection of 3H and 14C in polyacrylamide gels by fluorography. Eur J Biochem. 1975 Aug 15;56(2):335–341. doi: 10.1111/j.1432-1033.1975.tb02238.x. [DOI] [PubMed] [Google Scholar]
  19. Luckey M., Pollack J. R., Wayne R., Ames B. N., Neilands J. B. Iron uptake in Salmonella typhimurium: utilization of exogenous siderochromes as iron carriers. J Bacteriol. 1972 Sep;111(3):731–738. doi: 10.1128/jb.111.3.731-738.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. McIntosh M. A., Earhart C. F. Coordinate regulation by iron of the synthesis of phenolate compounds and three outer membrane proteins in Escherichia coli. J Bacteriol. 1977 Jul;131(1):331–339. doi: 10.1128/jb.131.1.331-339.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. McIntosh M. A., Earhart C. F. Effect of iron of the relative abundance of two large polypeptides of the Escherichia coli outer membrane. Biochem Biophys Res Commun. 1976 May 3;70(1):315–322. doi: 10.1016/0006-291x(76)91144-x. [DOI] [PubMed] [Google Scholar]
  22. O'Brien I. G., Gibson F. The structure of enterochelin and related 2,3-dihydroxy-N-benzoylserine conjugates from Escherichia coli. Biochim Biophys Acta. 1970 Aug 14;215(2):393–402. doi: 10.1016/0304-4165(70)90038-3. [DOI] [PubMed] [Google Scholar]
  23. Pollack J. R., Neilands J. B. Enterobactin, an iron transport compound from Salmonella typhimurium. Biochem Biophys Res Commun. 1970 Mar 12;38(5):989–992. doi: 10.1016/0006-291x(70)90819-3. [DOI] [PubMed] [Google Scholar]
  24. Pugsley A. P., Reeves P. Characterization of group B colicin-resistant mutants of Escherichia coli K-12: colicin resistance and the role of enterochelin. J Bacteriol. 1976 Jul;127(1):218–228. doi: 10.1128/jb.127.1.218-228.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Pugsley A. P., Reeves P. Increased production of the outer membrane receptors for colicins B, D and M by Escherichia coli under iron starvation. Biochem Biophys Res Commun. 1976 Jun 7;70(3):846–853. doi: 10.1016/0006-291x(76)90669-0. [DOI] [PubMed] [Google Scholar]
  26. Pugsley A. P., Reeves P. Iron uptake in colicin B-resistant mutants of Escherichia coli K-12. J Bacteriol. 1976 Jun;126(3):1052–1062. doi: 10.1128/jb.126.3.1052-1062.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Stiefel E. I., Watt G. D. Azotobacter cytochrome b557.5 is a bacterioferritin. Nature. 1979 May 3;279(5708):81–83. doi: 10.1038/279081a0. [DOI] [PubMed] [Google Scholar]
  28. Tait G. H. The identification and biosynthesis of siderochromes formed by Micrococcus denitrificans. Biochem J. 1975 Jan;146(1):191–204. doi: 10.1042/bj1460191. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Wayne R., Frick K., Neilands J. B. Siderophore protection against colicins M, B, V, and Ia in Escherichia coli. J Bacteriol. 1976 Apr;126(1):7–12. doi: 10.1128/jb.126.1.7-12.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Wayne R., Neilands J. B. Evidence for common binding sites for ferrichrome compounds and bacteriophage phi 80 in the cell envelope of Escherichia coli. J Bacteriol. 1975 Feb;121(2):497–503. doi: 10.1128/jb.121.2.497-503.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Worsham P. L., Konisky J. Use of cir-lac operon fusions to study transcriptional regulation of the colicin Ia receptor in Escherichia coli K-12. J Bacteriol. 1981 Jan;145(1):647–650. doi: 10.1128/jb.145.1.647-650.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Yagil G. Quantitative aspects of protein induction. Curr Top Cell Regul. 1975;9:183–236. doi: 10.1016/b978-0-12-152809-6.50013-5. [DOI] [PubMed] [Google Scholar]

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