Skip to main content
NCBI home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1973 Aug;115(2):615–622. doi: 10.1128/jb.115.2.615-622.1973

Derepression of Anthranilate Synthase in Purified Minicells of Escherichia coli Containing the Col-trp Plasmid

Anne Cornish Frazer a,1, Roy Curtiss III a,1
PMCID: PMC246290  PMID: 4579874

Abstract

Purified minicells of Escherichia coli K-12 containing the plasmid Col-trp+ or Col-trpA2 could be derepressed for the synthesis of anthranilate synthase, the first enzyme encoded in the trp operon. Non-plasmid-containing, deoxyribonucleic acid-deficient minicells could not be derepressed. Derepressed enzyme synthesis was initiated by l-tryptophan starvation. The kinetics of derepression were studied with minicells containing the Col-trpA2 plasmid. The derepression curves were biphasic with a rapid initial rate of enzyme synthesis followed by a slower rate of synthesis. The presence of l-tryptophan (20 to 50 μg/ml) or chloramphenicol (200 μg/ml) abolished enzyme synthesis. The presence of rifamycin SV (280 μg/ml) partially inhibited enzyme synthesis after at least 3.5 min of exposure. The ratio of minicell-to-cell synthetic capacity was 1:2.4 when compared on the basis of derepressed enzyme activity per unit cell volume. This work demonstrates that plasmid-containing minicells are capable of considerable functional protein and messenger ribonucleic acid synthesis and that the regulation of at least the trp operon is similar in minicells to that observed in cells.

Full text

PDF
615

Selected References

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

  1. Adler H. I., Fisher W. D., Cohen A., Hardigree A. A. MINIATURE escherichia coli CELLS DEFICIENT IN DNA. Proc Natl Acad Sci U S A. 1967 Feb;57(2):321–326. doi: 10.1073/pnas.57.2.321. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Baker R. F., Yanofsky C. The periodicity of RNA polymerase initiations: a new regulatory feature of transcription. Proc Natl Acad Sci U S A. 1968 May;60(1):313–320. doi: 10.1073/pnas.60.1.313. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Baker T. I., Crawford I. P. Anthranilate synthetase. Partial purification and some kinetic studies on the enzyme from Escherichia coli. J Biol Chem. 1966 Dec 10;241(23):5577–5584. [PubMed] [Google Scholar]
  4. CURTIS S. R., 3rd CHROMOSOMAL ABERRATIONS ASSOCIATED WITH MUTATIONS TO BACTERIOPHAGE RESISTANCE IN ESCHERICHIA COLI. J Bacteriol. 1965 Jan;89:28–40. doi: 10.1128/jb.89.1.28-40.1965. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cohen A., Fisher W. D., Curtiss R., 3rd, Adler H. I. The properties of DNA transferred to minicells during conjugation. Cold Spring Harb Symp Quant Biol. 1968;33:635–641. doi: 10.1101/sqb.1968.033.01.071. [DOI] [PubMed] [Google Scholar]
  6. Cohen S. N., Silver R. P., Sharp P. A., McCoubrey A. E. The problems of drug-resistant pathogenic bacteria. Studies on the molecular nature of R factors. Ann N Y Acad Sci. 1971 Jun 11;182:172–187. doi: 10.1111/j.1749-6632.1971.tb30655.x. [DOI] [PubMed] [Google Scholar]
  7. Curtiss R., 3rd, Charamella L. J., Stallions D. R., Mays J. A. Parental functions during conjugation in Escherichia coli K-12. Bacteriol Rev. 1968 Dec;32(4 Pt 1):320–348. [PMC free article] [PubMed] [Google Scholar]
  8. Fralick J. A., Fisher W. D., Adler H. I. Polyuridylic acid-directed phenylalanine incorporation in minicell extracts. J Bacteriol. 1969 Aug;99(2):621–622. doi: 10.1128/jb.99.2.621-622.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Gibson F. Chorismic acid: purification and some chemical and physical studies. Biochem J. 1964 Feb;90(2):256–261. doi: 10.1042/bj0900256. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Held W. A., Smith O. H. Mechanism of 3-methylanthranilic acid derepression of the tryptophan operon in Escherichia coli. J Bacteriol. 1970 Jan;101(1):209–217. doi: 10.1128/jb.101.1.209-217.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hickson F. T., Roth T. F., Helinski D. R. Circular DNA forms of a bacterial sex factor. Proc Natl Acad Sci U S A. 1967 Oct;58(4):1731–1738. doi: 10.1073/pnas.58.4.1731. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Inselburg J. R factor deoxyribonucleic acid in chromosomeless progeny of Escherichia coli. J Bacteriol. 1971 Feb;105(2):620–628. doi: 10.1128/jb.105.2.620-628.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Inselburg J. Segregation into and replication of plasmid deoxyribonucleic acid in chromosomeless segregants of Escherichia coli. J Bacteriol. 1970 Jun;102(3):642–647. doi: 10.1128/jb.102.3.642-647.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Ito J., Imamoto F. Sequential derepression and repression of the tryptophan operon in E. coli. Nature. 1968 Nov 2;220(5166):441–444. doi: 10.1038/220441a0. [DOI] [PubMed] [Google Scholar]
  15. Ito J., Yanofsky C. Anthranilate synthetase, an enzyme specified by the tryptophan operon of Escherichia coli: Comparative studies on the complex and the subunits. J Bacteriol. 1969 Feb;97(2):734–742. doi: 10.1128/jb.97.2.734-742.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kass L. R., Yarmolinsky M. B. Segregation of functional sex factor into minicells. Proc Natl Acad Sci U S A. 1970 Jul;66(3):815–822. doi: 10.1073/pnas.66.3.815. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kool A. J., Pranger M., Nijkamp H. J. Proteins synthesized by a non-induced bacteriocinogenic factor in minicells of Escherichia coli. Mol Gen Genet. 1972;115(4):314–323. doi: 10.1007/BF00333170. [DOI] [PubMed] [Google Scholar]
  18. LENNOX E. S. Transduction of linked genetic characters of the host by bacteriophage P1. Virology. 1955 Jul;1(2):190–206. doi: 10.1016/0042-6822(55)90016-7. [DOI] [PubMed] [Google Scholar]
  19. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  20. Levy S. B., Norman P. Segregation of transferable R factors into Escherichia coli minicells. Nature. 1970 Aug 8;227(5258):606–607. doi: 10.1038/227606a0. [DOI] [PubMed] [Google Scholar]
  21. Levy S. B. The problems of drug-resistant pathogenic bacteria. Studies on R factors segregated into E. coli minicells. Ann N Y Acad Sci. 1971 Jun 11;182:217–225. doi: 10.1111/j.1749-6632.1971.tb30658.x. [DOI] [PubMed] [Google Scholar]
  22. Morse D. E., Baker R. F., Yanofsky C. Translation of the tryptophan messenger RNA of Escherichia coli. Proc Natl Acad Sci U S A. 1968 Aug;60(4):1428–1435. doi: 10.1073/pnas.60.4.1428. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Roozen K. J., Fenwick R. G., Jr, Curtiss R., 3rd Synthesis of ribonucleic acid and protein in plasmid-containing minicells of Escherichia coli K-12. J Bacteriol. 1971 Jul;107(1):21–33. doi: 10.1128/jb.107.1.21-33.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. SCHAECHTER M., MAALOE O., KJELDGAARD N. O. Dependency on medium and temperature of cell size and chemical composition during balanced grown of Salmonella typhimurium. J Gen Microbiol. 1958 Dec;19(3):592–606. doi: 10.1099/00221287-19-3-592. [DOI] [PubMed] [Google Scholar]
  25. Wehrli W., Staehelin M. Actions of the rifamycins. Bacteriol Rev. 1971 Sep;35(3):290–309. doi: 10.1128/br.35.3.290-309.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES