Molecular and General Genetics MGG

, Volume 237, Issue 1–2, pp 129–133 | Cite as

DNA specificity of Escherichia coli deoP1 operator-DeoR repressor recognition

  • Karin Hammer
  • Lisbeth Bech
  • Palle Hobolth
  • Gert Dandanell
Article

Summary

We have studied the importance of the specific DNA sequence of the deo operator site for DeoR repressor binding by introducing symmetrical, single basepair substitutions at all positions in the deo operator and tested the ability of these variants to titrate DeoR in vivo. Our results show that a 16 by palindromic sequence constitutes the deo operator. Positions outside this palindrome (positions ±9, ±10) can be changed without any major effect on DeoR binding. Most of the central 6-8 by of the palindrome (positions ±1, ±2, ± 3) can be substituted with other nucleotides with no or only minor effects on DeoR binding, while changes at position ±4 and ±_5 give a more heterogeneous response. Finally, changes at positions ±6, ± 7 and ±8 severely disrupt DeoR binding.

Key words

DeoR repressor Repressor titration deo operator consensus sequence 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Amouyal M, Mortensen L, Buc H, Hammer K (1989) Single and double loop formation when deoR repressor binds to its natural operator sites. Cell 58:545–551Google Scholar
  2. Bech L (1991) Regulation of transcription initiation: Characterization of the DeoR repressor and DeoR-DNA interactions. PhD thesis, Uversity of CopenhagenGoogle Scholar
  3. Bremer E, Middendorf A, Martinussen J, Valentin-Hansen P (1990) Analysis of the tsx gene, which encodes a nucleoside-specific channel-forming protein (Tsx) in the outer membrane of Escherichia coli. Gene 96:59–65Google Scholar
  4. Buxton RS (1979) Fusion of the lac genes to the proximal promoters of the deo operon of Escherichia coli. J Gen Microbiol 112:241–250Google Scholar
  5. Clark DJ, Maaløoe O (1967) DNA replication and the division cycle of Escherichia coli. J Mol Biol 23:99–112Google Scholar
  6. Dandanell G (1992) DeoR repression at a distance only weakly responds to changes in interoperator separation and DNA topology. Nuc Acids Res 20:5407–5412Google Scholar
  7. Dandanell G, Hammer K (1985) Two operator sites separated by 599 base pairs are required for DeoR repression of the deo operon of Escherichia coli. EMBO J 4:3333–3338Google Scholar
  8. Dandanell G, Hammer K (1990) In vivo CRP-CAMP regulation of the deoP1 and deoP2 promoters of E. coli K12. Molecular Genetics (Live Sci Adv) 9:41–45Google Scholar
  9. Dandanell G, Hammer K (1991) deoP1 promoter and operator mutants in Escherichia coli: Isolation and characterization. Mol Microbiol 5:2371–2376Google Scholar
  10. Dandanell G, Norris K, Hammer K (1991) Long distance deoR regulation of gene expression in E. coli. Ann New York Acad Sci 646:19–30Google Scholar
  11. Dandanell G, Valentin-Hansen P, Løve-Larsen PE, Hammer K (1987) Long-range cooperativity between gene regulatory sequences in a prokaryote. Nature 325:823–826Google Scholar
  12. Davis T, Yamada M, Elgort M, Saier MH (1988) Nucleotide sequence of the mannitol (mtl) operon in Escherichia coli. Mol Microbiol 2:405–412Google Scholar
  13. Gicquel-Sanzey B, Cossart P (1982) Homologies between different procaryotic DNA-binding regulatory proteins and between their sites of action. EMBO J 1:591–595Google Scholar
  14. Hammer K, Dandanell G (1989) The deoR repressor from E. coli and its action in regulation-at-distance. In: Eckstein F, Lilley DMJ (eds) Nucleic acids and molecular biology. Springer-Verlag, Berlin- Heidelberg, pp 79–91Google Scholar
  15. Hammer-Jespersen K (1983) Nucleoside catabolism. In: MunchPetersen A (ed) Metabolism of nucleotides, nucleosides and nucleobases in microorganisms. Academic Press, London, pp 203–258Google Scholar
  16. Johnston F, Ponnambalam S, Busby S (1987) Binding of Escherichia coli RNA polymerase to a promoter carrying mutations that stop transcription initiation. J Mol Biol 195:745–748Google Scholar
  17. Koudelka GB, Harrison SC, Ptashne M (1987) Effect of non-contacted bases on the affinity of 434 operator for 434 repressor and cro. Nature 326:886–888Google Scholar
  18. Kriger-Brauer HJ, Braun V (1980) Functions related to the receptor protein specified by the tsx gene of Escherichia coli. Arch Microbiol 124:233–242Google Scholar
  19. Maniatis T, Fritsch EF, Sambrook J (1982) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New YorkGoogle Scholar
  20. Miller JH (1972) Experiments in molecular genetics. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New YorkGoogle Scholar
  21. Mortensen L, Dandanell G, Hammer K (1989) Purification and characterization of the DeoR repressor of Escherichia coli. EMBO J 8:325–331Google Scholar
  22. Munch-Petersen A, Jensen N (1990) Analysis of the regulatory region of the Escherichia coli nupG gene, encoding a nucleosidetransport protein. Eur J Biochem 190:547–551Google Scholar
  23. Munch-Petersen A, Mygind B (1983) Transport of nucleic acid precursors. In: Munch-Petersen A (ed) Metabolism of nucleotides, nucleosides and nucleobases in microorganisms. Academic Press, London, pp 259–305Google Scholar
  24. Munch-Petersen A, Nygaard P, Hammer-Jespersen K, Fiil N (1972) Mutants constitutive for nucleoside-catabolizing enzymes in Escherichia coli K12. Isolation, characterization and mapping. Eur J Biochem 27:208–215Google Scholar
  25. Oskouian B, Stewart GC (1990) Repression and catabolite repression of the lactose operon of Staphylococcus aureus. J Bacteriol 172:3804–3812Google Scholar
  26. O'Callaghan CH, Morris A, Kirby SM, Shingler AH (1972) Novel method for detection of β-lactamase by using a chromogenic cephalosporin substrate. Antimicrob Agents Chemother 1:283–288Google Scholar
  27. Pabo CO, Sauer RT (1984) Protein-DNA recognition. Ann Rev Biochem 53:293–321Google Scholar
  28. Ponnambalam S, Webster C, Birgham A, Busby S (1986) Transcription initiation of the Escherichia coli galactose operon promoters in the absence of normal — 35 regions sequences. J Biol Chem 261:16043–16048Google Scholar
  29. Rooijen RJ van, Vos WM de (1990) Molecular cloning, transcriptional analysis and nucleotide sequence of lacR, a gene encoding the repressor of the lactose phosphotransferase system of Lactococcus lactis. J Biol Chem 165:18499–18503Google Scholar
  30. Silhavy TJ, Berman ML, Enquist LW (1984) Experiments with gene fusions. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New YorkGoogle Scholar
  31. Valentin-Hansen P, Aiba H, Schümperli D (1982) The structure of tandem regulatory regions in the deo operon of Escherichia coli K12. EMBO J 1:317–322Google Scholar
  32. Valentin-Hansen P, Albrechtsen B, Larsen JEL (1986) DNA-protein recognition: demonstration of three genetically separated operator elements that are required for the repression of the Escherichia coli deoCABD promoters by the DeoR repressor. EMBO J 5:2015–2021Google Scholar

Copyright information

© Springer-Verlag 1993

Authors and Affiliations

  • Karin Hammer
    • 1
  • Lisbeth Bech
    • 2
  • Palle Hobolth
    • 3
  • Gert Dandanell
    • 4
  1. 1.Department of MicrobiologyTechnical University of DenmarkLyngby, CopenhagenDenmark
  2. 2.Novo Research InstituteBagsærdDenmark
  3. 3.Pharmacia LKBHillerodDenmark
  4. 4.Institute of Biological Chemistry BUniversity of CopenhagenCopenhagen KDenmark

Personalised recommendations