Skip to main content
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1985 Jun 11;13(11):4113–4123. doi: 10.1093/nar/13.11.4113

Codon-defined ribosomal pausing in Escherichia coli detected by using the pyrE attenuator to probe the coupling between transcription and translation.

F Bonekamp, H D Andersen, T Christensen, K F Jensen
PMCID: PMC341299  PMID: 2989788

Abstract

This communication describes an assay for the relative translation efficiency of individual codons which makes use of the pyrE attenuator to probe the coupling between transcription and translation at the end of an artificial leader peptide. By cloning of short synthetic DNA fragments the codons to be tested were placed in the middle of the leader peptide and the downstream transcription of a pyrE"lacZ gene was monitored by measuring beta-galactosidase activity. The substitution, one by one, of three AGG codons for arginine with three CGT codons for the same amino acid residue was found to cause a two fold increase per codon of transcription over the pyrE attenuator, such that an eight fold higher frequency of pyrE expression was seen when all three AGG codons were replaced by CGT codons. No such effect of codon composition was observed, when the cells were grown with a low UTP pool which causes a reduction of the mRNA chain growth rate.

Full text

PDF
4113

Selected References

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

  1. Adhya S., Gottesman M. Control of transcription termination. Annu Rev Biochem. 1978;47:967–996. doi: 10.1146/annurev.bi.47.070178.004535. [DOI] [PubMed] [Google Scholar]
  2. Bonekamp F., Clemmesen K., Karlström O., Jensen K. F. Mechanism of UTP-modulated attenuation at the pyrE gene of Escherichia coli: an example of operon polarity control through the coupling of translation to transcription. EMBO J. 1984 Dec 1;3(12):2857–2861. doi: 10.1002/j.1460-2075.1984.tb02220.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Gouy M., Gautier C. Codon usage in bacteria: correlation with gene expressivity. Nucleic Acids Res. 1982 Nov 25;10(22):7055–7074. doi: 10.1093/nar/10.22.7055. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Grantham R., Gautier C., Gouy M., Jacobzone M., Mercier R. Codon catalog usage is a genome strategy modulated for gene expressivity. Nucleic Acids Res. 1981 Jan 10;9(1):r43–r74. doi: 10.1093/nar/9.1.213-b. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Grosjean H., Fiers W. Preferential codon usage in prokaryotic genes: the optimal codon-anticodon interaction energy and the selective codon usage in efficiently expressed genes. Gene. 1982 Jun;18(3):199–209. doi: 10.1016/0378-1119(82)90157-3. [DOI] [PubMed] [Google Scholar]
  6. Ikemura T. Correlation between the abundance of Escherichia coli transfer RNAs and the occurrence of the respective codons in its protein genes. J Mol Biol. 1981 Feb 15;146(1):1–21. doi: 10.1016/0022-2836(81)90363-6. [DOI] [PubMed] [Google Scholar]
  7. Ikemura T. Correlation between the abundance of Escherichia coli transfer RNAs and the occurrence of the respective codons in its protein genes: a proposal for a synonymous codon choice that is optimal for the E. coli translational system. J Mol Biol. 1981 Sep 25;151(3):389–409. doi: 10.1016/0022-2836(81)90003-6. [DOI] [PubMed] [Google Scholar]
  8. Jensen K. F., Larsen J. N., Schack L., Sivertsen A. Studies on the structure and expression of Escherichia coli pyrC, pyrD, and pyrF using the cloned genes. Eur J Biochem. 1984 Apr 16;140(2):343–352. doi: 10.1111/j.1432-1033.1984.tb08107.x. [DOI] [PubMed] [Google Scholar]
  9. Jensen K. F., Neuhard J., Schack L. RNA polymerase involvement in the regulation of expression of Salmonella typhimurium pyr genes. Isolation and characterization of a fluorouracil-resistant mutant with high, constitutive expression of the pyrB and pyrE genes due to a mutation in rpoBC. EMBO J. 1982;1(1):69–74. doi: 10.1002/j.1460-2075.1982.tb01126.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Konigsberg W., Godson G. N. Evidence for use of rare codons in the dnaG gene and other regulatory genes of Escherichia coli. Proc Natl Acad Sci U S A. 1983 Feb;80(3):687–691. doi: 10.1073/pnas.80.3.687. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Pedersen S. Escherichia coli ribosomes translate in vivo with variable rate. EMBO J. 1984 Dec 1;3(12):2895–2898. doi: 10.1002/j.1460-2075.1984.tb02227.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Poulsen P., Bonekamp F., Jensen K. F. Structure of the Escherichia coli pyrE operon and control of pyrE expression by a UTP modulated intercistronic attentuation. EMBO J. 1984 Aug;3(8):1783–1790. doi: 10.1002/j.1460-2075.1984.tb02046.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Poulsen P., Jensen K. F., Valentin-Hansen P., Carlsson P., Lundberg L. G. Nucleotide sequence of the Escherichia coli pyrE gene and of the DNA in front of the protein-coding region. Eur J Biochem. 1983 Sep 15;135(2):223–229. doi: 10.1111/j.1432-1033.1983.tb07641.x. [DOI] [PubMed] [Google Scholar]
  14. Robinson M., Lilley R., Little S., Emtage J. S., Yarranton G., Stephens P., Millican A., Eaton M., Humphreys G. Codon usage can affect efficiency of translation of genes in Escherichia coli. Nucleic Acids Res. 1984 Sep 11;12(17):6663–6671. doi: 10.1093/nar/12.17.6663. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Sanchez-Pescador R., Urdea M. S. Use of unpurified synthetic deoxynucleotide primers for rapid dideoxynucleotide chain termination sequencing. DNA. 1984 Aug;3(4):339–343. doi: 10.1089/dna.1.1984.3.339. [DOI] [PubMed] [Google Scholar]
  16. 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]
  17. Turnbough C. L., Jr, Hicks K. L., Donahue J. P. Attenuation control of pyrBI operon expression in Escherichia coli K-12. Proc Natl Acad Sci U S A. 1983 Jan;80(2):368–372. doi: 10.1073/pnas.80.2.368. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES