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. 1974 Jan;71(1):11–15. doi: 10.1073/pnas.71.1.11

Synthesis of Diphtheria tox-Gene Products in Escherichia coli Extracts

J R Murphy 1, A M Pappenheimer Jr 1, S Tayart De Borms 1
PMCID: PMC387921  PMID: 4204202

Abstract

In a protein-synthesizing system extracted from E. coli, purified DNA from corynephages βctox+ and β45c was used to direct the in vitro synthesis of diphtheria toxin and of the related nontoxic protein, CRM45, as well as of other β-phage proteins. When βctox+-DNA or βc-DNA was added to a similar system extracted from the nonlysogenic Corynebacterium diphtheriae strain, C7s(-)tox-, neither toxin nor the CRM45 protein was produced, although other β-phage proteins were synthesized in amounts equivalent to those produced in the E. coli system from the same amount of β-phage DNA. Preliminary experiments suggest that both toxinogenic and nontoxinogenic strains of the diphtheria bacillus contain a factor that specificially blocks expression of the tox gene. Synthesis of toxin and the CRM45 protein in the E. coli system could not be inhibited by relatively high concentrations of inorganic iron, but could be inhibited by extracts from the C7s(-)tox- strain of C. diphtheriae.

Keywords: diphtheria toxin, in vitro protein synthesis, corynephage β

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Selected References

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  1. Drazin R., Kandel J., Collier R. J. Structure and activity of diphtheria toxin. II. Attack by trypsin at a specific site within the intact toxin molecule. J Biol Chem. 1971 Mar 10;246(5):1504–1510. [PubMed] [Google Scholar]
  2. FREEMAN V. J. Studies on the virulence of bacteriophage-infected strains of Corynebacterium diphtheriae. J Bacteriol. 1951 Jun;61(6):675–688. doi: 10.1128/jb.61.6.675-688.1951. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Gesteland R. F., Kahn C. Synthesis of bacteriophage lambda proteins in vitro. Nat New Biol. 1972 Nov 1;240(96):3–6. doi: 10.1038/newbio240003a0. [DOI] [PubMed] [Google Scholar]
  4. Gill D. M., Dinius L. L. Observations on the structure of diphtheria toxin. J Biol Chem. 1971 Mar 10;246(5):1485–1491. [PubMed] [Google Scholar]
  5. Gill D. M., Pappenheimer A. M., Jr Structure-activity relationships in diphtheria toxin. J Biol Chem. 1971 Mar 10;246(5):1492–1495. [PubMed] [Google Scholar]
  6. Gill D. M., Uchida T., Singer R. A. Expression of diphtheria toxin genes carried by integrated and nonintegrated phage beta. Virology. 1972 Dec;50(3):664–668. doi: 10.1016/0042-6822(72)90420-5. [DOI] [PubMed] [Google Scholar]
  7. Gold L. M., Schweiger M. Synthesis of phage-specific alpha- and beta-glucosyl transferases directed by T-even DNA in vitro. Proc Natl Acad Sci U S A. 1969 Mar;62(3):892–898. doi: 10.1073/pnas.62.3.892. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Greenblatt J. Positive control of endolysin synthesis in vitro by the gene N protein of phage lambda. Proc Natl Acad Sci U S A. 1972 Dec;69(12):3606–3610. doi: 10.1073/pnas.69.12.3606. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. HATANO M. Effect of iron concentration in the medium on phage and toxin production in a lysogenic, virulent Corynebacterium diphtheriae. J Bacteriol. 1956 Jan;71(1):121–122. doi: 10.1128/jb.71.1.121-122.1956. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Matsuda M., Barksdale L. System for the investigation of the bacteriophage-directed synthesis of diphtherial toxin. J Bacteriol. 1967 Feb;93(2):722–730. doi: 10.1128/jb.93.2.722-730.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Pappenheimer A. M., Jr, Gill D. M. Diphtheria. Science. 1973 Oct 26;182(4110):353–358. doi: 10.1126/science.182.4110.353. [DOI] [PubMed] [Google Scholar]
  12. RAYNAUD M., RELYVELD E. H. [Diphtheria toxin-antitoxin reaction]. Ann Inst Pasteur (Paris) 1959 Nov;97:636–678. [PubMed] [Google Scholar]
  13. Richter D., Herrlich P., Schweiger M. Phage DNA directed enzyme synthesis in vitro system from yeast mitochondria. Nat New Biol. 1972 Jul 19;238(81):74–76. doi: 10.1038/newbio238074a0. [DOI] [PubMed] [Google Scholar]
  14. Uchida T., Gill D. M., Pappenheimer A. M., Jr Mutation in the structural gene for diphtheria toxin carried by temperate phage . Nat New Biol. 1971 Sep 1;233(35):8–11. doi: 10.1038/newbio233008a0. [DOI] [PubMed] [Google Scholar]
  15. Uchida T., Pappenheimer A. M., Jr, Greany R. Diphtheria toxin and related proteins. I. Isolation and properties of mutant proteins serologically related to diphtheria toxin. J Biol Chem. 1973 Jun 10;248(11):3838–3844. [PubMed] [Google Scholar]
  16. YONEDA M., PAPPENHEIMER A. M., Jr Some effects of iron deficiency on the extracellular products released by toxigenic and nontoxigenic strains of Corynebacterium diphtheriae. J Bacteriol. 1957 Aug;74(2):256–264. doi: 10.1128/jb.74.2.256-264.1957. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. 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]

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