Skip to main content
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1996 Jul 15;24(14):2712–2717. doi: 10.1093/nar/24.14.2712

Site-specific recombination by the beta protein from the streptococcal plasmid pSM19035: minimal recombination sequences and crossing over site.

I Canosa 1, F Rojo 1, J C Alonso 1
PMCID: PMC146011  PMID: 8759001

Abstract

The beta recombinase from the broad host range Grampositive plasmid pSM19035 catalyzes intramolecular site-specific recombination between two directly or inversely oriented recombination sites in the presence of a chromatin-associated protein (Hbsu). The recombination site had been localized to a 447 bp DNA segment from pSM19035. This segment includes a 90 bp region that contains two adjacent binding sites (I and II) for beta protein dimers. Using in vitro recombination assays, we show that this 90 bp region is necessary and sufficient for beta protein-mediated recombination; this defines the six site as the region required for beta protein binding. The point of crossing over has been localized to the center of site I. Hbsu has a strong binding affinity for an unknown site located within the 447 bp segment containing the six site. We discuss the possibility that Hbsu recognizes an altered DNA structure, rather than a specific sequence, generated in the synaptic complex.

Full Text

The Full Text of this article is available as a PDF (91.8 KB).

Selected References

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

  1. Alonso J. C., Gutierrez C., Rojo F. The role of chromatin-associated protein Hbsu in beta-mediated DNA recombination is to facilitate the joining of distant recombination sites. Mol Microbiol. 1995 Nov;18(3):471–478. doi: 10.1111/j.1365-2958.1995.mmi_18030471.x. [DOI] [PubMed] [Google Scholar]
  2. Alonso J. C., Weise F., Rojo F. The Bacillus subtilis histone-like protein Hbsu is required for DNA resolution and DNA inversion mediated by the beta recombinase of plasmid pSM19035. J Biol Chem. 1995 Feb 17;270(7):2938–2945. doi: 10.1074/jbc.270.7.2938. [DOI] [PubMed] [Google Scholar]
  3. Bednarz A. L., Boocock M. R., Sherratt D. J. Determinants of correct res site alignment in site-specific recombination by Tn3 resolvase. Genes Dev. 1990 Dec;4(12B):2366–2375. doi: 10.1101/gad.4.12b.2366. [DOI] [PubMed] [Google Scholar]
  4. Bonnefoy E., Takahashi M., Yaniv J. R. DNA-binding parameters of the HU protein of Escherichia coli to cruciform DNA. J Mol Biol. 1994 Sep 16;242(2):116–129. doi: 10.1006/jmbi.1994.1563. [DOI] [PubMed] [Google Scholar]
  5. Cegłowski P., Alonso J. C. Gene organization of the Streptococcus pyogenes plasmid pDB101: sequence analysis of the orf eta-copS region. Gene. 1994 Jul 22;145(1):33–39. doi: 10.1016/0378-1119(94)90319-0. [DOI] [PubMed] [Google Scholar]
  6. Cegłowski P., Boitsov A., Chai S., Alonso J. C. Analysis of the stabilization system of pSM19035-derived plasmid pBT233 in Bacillus subtilis. Gene. 1993 Dec 22;136(1-2):1–12. doi: 10.1016/0378-1119(93)90441-5. [DOI] [PubMed] [Google Scholar]
  7. Grindley N. D., Lauth M. R., Wells R. G., Wityk R. J., Salvo J. J., Reed R. R. Transposon-mediated site-specific recombination: identification of three binding sites for resolvase at the res sites of gamma delta and Tn3. Cell. 1982 Aug;30(1):19–27. doi: 10.1016/0092-8674(82)90007-1. [DOI] [PubMed] [Google Scholar]
  8. Haykinson M. J., Johnson R. C. DNA looping and the helical repeat in vitro and in vivo: effect of HU protein and enhancer location on Hin invertasome assembly. EMBO J. 1993 Jun;12(6):2503–2512. doi: 10.1002/j.1460-2075.1993.tb05905.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Johnson R. C. Mechanism of site-specific DNA inversion in bacteria. Curr Opin Genet Dev. 1991 Oct;1(3):404–411. doi: 10.1016/s0959-437x(05)80307-7. [DOI] [PubMed] [Google Scholar]
  10. Lavoie B. D., Chaconas G. A second high affinity HU binding site in the phage Mu transpososome. J Biol Chem. 1994 Jun 3;269(22):15571–15576. [PubMed] [Google Scholar]
  11. Lavoie B. D., Chaconas G. Site-specific HU binding in the Mu transpososome: conversion of a sequence-independent DNA-binding protein into a chemical nuclease. Genes Dev. 1993 Dec;7(12B):2510–2519. doi: 10.1101/gad.7.12b.2510. [DOI] [PubMed] [Google Scholar]
  12. Leschziner A. E., Boocock M. R., Grindley N. D. The tyrosine-6 hydroxyl of gamma delta resolvase is not required for the DNA cleavage and rejoining reactions. Mol Microbiol. 1995 Mar;15(5):865–870. doi: 10.1111/j.1365-2958.1995.tb02356.x. [DOI] [PubMed] [Google Scholar]
  13. Paull T. T., Haykinson M. J., Johnson R. C. The nonspecific DNA-binding and -bending proteins HMG1 and HMG2 promote the assembly of complex nucleoprotein structures. Genes Dev. 1993 Aug;7(8):1521–1534. doi: 10.1101/gad.7.8.1521. [DOI] [PubMed] [Google Scholar]
  14. Petit M. A., Ehrlich D., Jannière L. pAM beta 1 resolvase has an atypical recombination site and requires a histone-like protein HU. Mol Microbiol. 1995 Oct;18(2):271–282. doi: 10.1111/j.1365-2958.1995.mmi_18020271.x. [DOI] [PubMed] [Google Scholar]
  15. Pontiggia A., Negri A., Beltrame M., Bianchi M. E. Protein HU binds specifically to kinked DNA. Mol Microbiol. 1993 Feb;7(3):343–350. doi: 10.1111/j.1365-2958.1993.tb01126.x. [DOI] [PubMed] [Google Scholar]
  16. Pujol C., Ehrlich S. D., Jannière L. The promiscuous plasmids pIP501 and pAM beta 1 from gram-positive bacteria encode complementary resolution functions. Plasmid. 1994 Jan;31(1):100–105. doi: 10.1006/plas.1994.1010. [DOI] [PubMed] [Google Scholar]
  17. Reed R. R., Grindley N. D. Transposon-mediated site-specific recombination in vitro: DNA cleavage and protein-DNA linkage at the recombination site. Cell. 1981 Sep;25(3):721–728. doi: 10.1016/0092-8674(81)90179-3. [DOI] [PubMed] [Google Scholar]
  18. Rojo F., Alonso J. C. A novel site-specific recombinase encoded by the Streptococcus pyogenes plasmid pSM19035. J Mol Biol. 1994 Apr 29;238(2):159–172. doi: 10.1006/jmbi.1994.1278. [DOI] [PubMed] [Google Scholar]
  19. Rojo F., Alonso J. C. The beta recombinase from the Streptococcal plasmid pSM 19035 represses its own transcription by holding the RNA polymerase at the promoter region. Nucleic Acids Res. 1994 May 25;22(10):1855–1860. doi: 10.1093/nar/22.10.1855. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Rojo F., Alonso J. C. The beta recombinase of plasmid pSM19035 binds to two adjacent sites, making different contacts at each of them. Nucleic Acids Res. 1995 Aug 25;23(16):3181–3188. doi: 10.1093/nar/23.16.3181. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Rojo F., Weise F., Alonso J. C. Purification of the beta product encoded by the Streptococcus pyogenes plasmid pSM19035. A putative DNA recombinase required to resolve plasmid oligomers. FEBS Lett. 1993 Aug 9;328(1-2):169–173. doi: 10.1016/0014-5793(93)80987-6. [DOI] [PubMed] [Google Scholar]
  22. Segall A. M., Goodman S. D., Nash H. A. Architectural elements in nucleoprotein complexes: interchangeability of specific and non-specific DNA binding proteins. EMBO J. 1994 Oct 3;13(19):4536–4548. doi: 10.1002/j.1460-2075.1994.tb06775.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Stark W. M., Boocock M. R., Sherratt D. J. Catalysis by site-specific recombinases. Trends Genet. 1992 Dec;8(12):432–439. [PubMed] [Google Scholar]
  24. Stark W. M., Grindley N. D., Hatfull G. F., Boocock M. R. Resolvase-catalysed reactions between res sites differing in the central dinucleotide of subsite I. EMBO J. 1991 Nov;10(11):3541–3548. doi: 10.1002/j.1460-2075.1991.tb04918.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. van de Putte P., Goosen N. DNA inversions in phages and bacteria. Trends Genet. 1992 Dec;8(12):457–462. doi: 10.1016/0168-9525(92)90331-w. [DOI] [PubMed] [Google Scholar]

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

RESOURCES