Mutational analysis of a prokaryotic recombinational enhancer element with two functions - PubMed (original) (raw)

Mutational analysis of a prokaryotic recombinational enhancer element with two functions

P Hübner et al. EMBO J. 1989 Feb.

Abstract

The site-specific DNA inversion system Cin encoded by the bacteriophage P1 consists of a recombinase, two inverted crossing-over sites and a recombinational enhancer. The latter approximately 75 bp long genetic element is bifunctional due to its location within the 5' part of the cin gene encoding the recombinase. In order to determine the essential nucleotides for each of its two biological functions we randomly mutated the recombinational enhancer sequence sis(P1) and analysed both functions of the mutants obtained. Three distinct regions of this sequence were found to be important for the enhancer activity. One of them occupies the middle third of the enhancer sequence and it can suffer a number of functionally neutral base substitutions, while others are detrimental. The other two regions occupy the two flanking thirds of the enhancer. They coincide with binding sites of the host-coded protein FIS (Factor for Inversion Stimulation) needed for efficient DNA inversion in vitro. These sequences appear to be highly evolved allowing only a few mutations without affecting either of the biological functions. Taking the effect of mutations within these FIS binding sites into account a consensus sequence for the interaction with FIS was compiled. This FIS consensus implies a palindromic structure for the recombinational enhancer. This is in line with the orientation independence of enhancer action with respect to the crossing-over sites.

PubMed Disclaimer

Similar articles

• Bent DNA is needed for recombinational enhancer activity in the site-specific recombination system Cin of bacteriophage P1. The role of FIS protein.
  Hübner P, Haffter P, Iida S, Arber W. Hübner P, et al. J Mol Biol. 1989 Feb 5;205(3):493-500. doi: 10.1016/0022-2836(89)90220-9. J Mol Biol. 1989. PMID: 2648006
• Purification and DNA-binding properties of FIS and Cin, two proteins required for the bacteriophage P1 site-specific recombination system, cin.
  Haffter P, Bickle TA. Haffter P, et al. J Mol Biol. 1987 Dec 20;198(4):579-87. doi: 10.1016/0022-2836(87)90201-4. J Mol Biol. 1987. PMID: 3323534
• The recombinational enhancer for DNA inversion functions independent of its orientation as a consequence of dyad symmetry in the Fis-DNA complex.
  Kanaar R, van Hal JP, van de Putte P. Kanaar R, et al. Nucleic Acids Res. 1989 Aug 11;17(15):6043-53. doi: 10.1093/nar/17.15.6043. Nucleic Acids Res. 1989. PMID: 2549506 Free PMC article.
• DNA inversions in phages and bacteria.
  van de Putte P, Goosen N. van de Putte P, et al. Trends Genet. 1992 Dec;8(12):457-62. doi: 10.1016/0168-9525(92)90331-w. Trends Genet. 1992. PMID: 1337227 Review.
• Structure and function of the shufflon in plasmid R64.
  Gyohda A, Furuya N, Ishiwa A, Zhu S, Komano T. Gyohda A, et al. Adv Biophys. 2004;38:183-213. Adv Biophys. 2004. PMID: 15493334 Review.

Cited by

• The N-terminal part of the E.coli DNA binding protein FIS is essential for stimulating site-specific DNA inversion but is not required for specific DNA binding.
  Koch C, Ninnemann O, Fuss H, Kahmann R. Koch C, et al. Nucleic Acids Res. 1991 Nov 11;19(21):5915-22. doi: 10.1093/nar/19.21.5915. Nucleic Acids Res. 1991. PMID: 1834996 Free PMC article.
• The shape of the DNA minor groove directs binding by the DNA-bending protein Fis.
  Stella S, Cascio D, Johnson RC. Stella S, et al. Genes Dev. 2010 Apr 15;24(8):814-26. doi: 10.1101/gad.1900610. Genes Dev. 2010. PMID: 20395367 Free PMC article.
• Genome of bacteriophage P1.
  Łobocka MB, Rose DJ, Plunkett G 3rd, Rusin M, Samojedny A, Lehnherr H, Yarmolinsky MB, Blattner FR. Łobocka MB, et al. J Bacteriol. 2004 Nov;186(21):7032-68. doi: 10.1128/JB.186.21.7032-7068.2004. J Bacteriol. 2004. PMID: 15489417 Free PMC article.
• The Escherichia coli gapA gene is transcribed by the vegetative RNA polymerase holoenzyme E sigma 70 and by the heat shock RNA polymerase E sigma 32.
  Charpentier B, Branlant C. Charpentier B, et al. J Bacteriol. 1994 Feb;176(3):830-9. doi: 10.1128/jb.176.3.830-839.1994. J Bacteriol. 1994. PMID: 8300536 Free PMC article.
• Location, degree, and direction of DNA bending associated with the Hin recombinational enhancer sequence and Fis-enhancer complex.
  Perkins-Balding D, Dias DP, Glasgow AC. Perkins-Balding D, et al. J Bacteriol. 1997 Aug;179(15):4747-53. doi: 10.1128/jb.179.15.4747-4753.1997. J Bacteriol. 1997. PMID: 9244261 Free PMC article.

References

1. 1. Biochim Biophys Acta. 1984 Jun 16;782(2):111-9 - PubMed
2. 1. Proc Natl Acad Sci U S A. 1985 Jun;82(11):3776-80 - PubMed
3. 1. EMBO J. 1983;2(10):1733-40 - PubMed
4. 1. J Gen Virol. 1986 Jun;67 ( Pt 6):1123-33 - PubMed
5. 1. EMBO J. 1988 Apr;7(4):1229-37 - PubMed

Publication types

MeSH terms

Substances

LinkOut - more resources

• Full Text Sources

  • Europe PubMed Central
  • Nature Publishing Group
  • PubMed Central
  • Wiley

• Research Materials

  • NCI CPTC Antibody Characterization Program