Understanding DNA Looping Through Cre-Recombination Kinetics

  • Massa J. ShouraEmail author
  • Stephen D. Levene
Part of the Natural Computing Series book series (NCS)


The interior of a cell is a crowded and fluctuating environment where DNA and other biomolecules are both highly constrained and subject to many mechanical forces. The extensive compaction of DNA in living cells is a challenge to many critical biological functions. An evolutionary solution to this challenge may be the juxtaposition of cis-acting elements such that multimeric protein complexes simultaneously interact with two or more protein-binding sites. This mode of biological activity involves the formation of looped DNA structures, which, by themselves, are thermodynamically unfavorable. Our knowledge about the roles of DNA bending, twisting, and their respective energetics in DNA looping has come mainly from analyses of ligase-dependent DNA cyclization experiments, which are quantitatively described by the Jacobson–Stockmayer, or J, factor. In this chapter, we discuss a novel quantitative approach to measuring the probability of DNA loop formation in solution using ensemble Förster resonance energy transfer (FRET) measurements of intramolecular and intermolecular Cre-recombination kinetics. Because the mechanism of Cre recombinase does not conform to a simple kinetic scheme, we employ numerical methods to extract rate constants for fundamental steps that pertain to Cre-mediated loop closure.


Extract Rate Constant Intermolecular Reaction Transcription Factor Interaction Intramolecular Recombination Repeat loxP Site 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



We thank Andreas Hanke and Stefan Giovan for calculations of J theor. This work was supported by a grant from the NIH/NSF Joint Program in Mathematical Biology (DMS-0800929 from the National Science Foundation) to SDL.


  1. 1.
    Y. Zhang, D.M. Crothers, Proc. Natl. Acad. Sci. U. S. A. 100, 3161–3166 (2003)CrossRefGoogle Scholar
  2. 2.
    T. Förster, Z. Naturforsch. A. 4(7) (1949)Google Scholar
  3. 3.
    R.E. Dale, J. Eisinger, Biopolymers 13, 1573–1605 (1974)CrossRefGoogle Scholar
  4. 4.
    R.E. Dale, J. Eisinger, W.E. Blumberg, Biophys. J. 26, 161–193 (1979)CrossRefGoogle Scholar
  5. 5.
    D. Badali, C.C. Gradinaru, J. Chem. Phys. 134, 225102 (2011)CrossRefGoogle Scholar
  6. 6.
    R.D. Mitra, C.M. Silva, D.C. Youvan, Gene 173, 13–17 (1996)CrossRefGoogle Scholar
  7. 7.
    R. Day, Mol. Endocrinol. 12, 1410–1419 (1998)CrossRefGoogle Scholar
  8. 8.
    B. Treutlein, A. Muschielok, J. Andrecka, A. Jawhari, C. Buchen, D. Kostrewa, F. Hog, P. Cramer, J. Michaelis, Mol. Cell 46, 136–146 (2012)CrossRefGoogle Scholar
  9. 9.
    S.M. Miick, R.S. Fee, D.P. Millar, W.J. Chazin, Proc. Natl. Acad. Sci. U. S. A. 94, 9080–9084 (1997)CrossRefGoogle Scholar
  10. 10.
    A.K. Wozniak, G.F. Schroder, H. Grubmuller, C.A. Seidel, F. Oesterhelt, Proc. Natl. Acad. Sci. U. S. A. 105, 18337–18342 (2008)CrossRefGoogle Scholar
  11. 11.
    S.A. McKinney, A.D. Freeman, D.M. Lilley, T. Ha, Proc. Natl. Acad. Sci. U. S. A. 102, 5715–5720 (2005)CrossRefGoogle Scholar
  12. 12.
    E. Tan, T.J. Wilson, M.K. Nahas, R.M. Clegg, D.M. Lilley, T. Ha, Proc. Natl. Acad. Sci. U. S. A.100, 9308–9313 (2003)CrossRefGoogle Scholar
  13. 13.
    M. Bussiek, K. Toth, N. Schwarz, J. Langowski, Biochemistry 45, 10838–10846 (2006)CrossRefGoogle Scholar
  14. 14.
    C.L. White, K. Luger, J. Mol. Biol. 342, 1391–1402 (2004)CrossRefGoogle Scholar
  15. 15.
    M. Tomschik, K. van Holde, J. Zlatanova, J. Fluoresc. 19, 53–62 (2009)CrossRefGoogle Scholar
  16. 16.
    C. Bonisch, K. Schneider, S. Punzeler, S.M. Wiedemann, C. Bielmeier, M. Bocola, H.C. Eberl, W. Kuegel, J. Neumann, E. Kremmer, H. Leonhardt, M. Mann, J. Michaelis, L. Schermelleh, S.B. Hake, Nucleic Acids Res. 40, 5951–5964 (2012)CrossRefGoogle Scholar
  17. 17.
    R. Zhou, T. Ha, Methods Mol. Biol. 922, 85–100 (2012)Google Scholar
  18. 18.
    S.F. Singleton, J. Xiao, Biopolymers 61, 145–158 (2001)CrossRefGoogle Scholar
  19. 19.
    M. Margittai, J. Widengren, E. Schweinberger, G.F. Schroder, S. Felekyan, E. Haustein, M. Konig, D. Fasshauer, H. Grubmuller, R. Jahn, C.A. Seidel, Proc. Natl. Acad. Sci. U. S. A. 100, 15516–15521 (2003)CrossRefGoogle Scholar
  20. 20.
    T. Ha, A.G. Kozlov, T.M. Lohman, Annu. Rev. Biophys. 41, 295–319 (2012)CrossRefGoogle Scholar
  21. 21.
    W.M. Stark, D.J. Sherratt, M.R. Boocock, Cell 58, 779–790 (1989)CrossRefGoogle Scholar
  22. 22.
    S.M. Lewis, Adv. Immunol. 56, 27–150 (1994)CrossRefGoogle Scholar
  23. 23.
    D.J. Sherratt, L.K. Arciszewska, G. Blakely, S. Colloms, K. Grant, N. Leslie, R. McCulloch, Philos. Trans. R. Soc. Lond. B Biol. Sci. 347, 37–42 (1995)CrossRefGoogle Scholar
  24. 24.
    B. Hallet, D.J. Sherratt, FEMS Microbiol. Rev. 21, 157–178 (1997)CrossRefGoogle Scholar
  25. 25.
    R. Oh-McGinnis, M.J. Jones, L. Lefebvre, Brief. Funct. Genomics 9, 281–293 (2010)CrossRefGoogle Scholar
  26. 26.
    A. Landy, Annu. Rev. Biochem. 58, 913–949 (1989)CrossRefGoogle Scholar
  27. 27.
    J.R. Scott, Virology 36, 564–574 (1968)CrossRefGoogle Scholar
  28. 28.
    N. Sternberg, Cold Spring Harb. Symp. Quant. Biol. 43(Pt 2), 1143–1146 (1979)CrossRefMathSciNetGoogle Scholar
  29. 29.
    N. Sternberg, D. Hamilton, S. Austin, M. Yarmolinsky, R. Hoess, Cold Spring Harb. Symp. Quant. Biol. 45(Pt 1), 297–309 (1981)CrossRefGoogle Scholar
  30. 30.
    C.H. Ma, A.H. Kachroo, A. Macieszak, T.Y. Chen, P. Guga, M. Jayaram, PLoS One 4, e7248 (2009)CrossRefGoogle Scholar
  31. 31.
    D.N. Gopaul, F. Guo, G.D. Van Duyne, EMBO J 17, 4175–4187 (1998)CrossRefGoogle Scholar
  32. 32.
    K. Ghosh, G.D. Van Duyne, Methods 28, 374–383 (2002)CrossRefGoogle Scholar
  33. 33.
    A.A. Vetcher, A.Y. Lushnikov, J. Navarra-Madsen, R.G. Scharein, Y.L. Lyubchenko, I.K. Darcy, S.D. Levene, J. Mol. Biol. 357, 1089–1104 (2006)CrossRefGoogle Scholar
  34. 34.
    F. Guo, D.N. Gopaul, G.D. van Duyne, Nature 389, 40–46 (1997)CrossRefGoogle Scholar
  35. 35.
    F. Guo, D.N. Gopaul, G.D. Van Duyne, Proc. Natl. Acad. Sci. U. S. A. 96, 7143–7148 (1999)CrossRefGoogle Scholar
  36. 36.
    E. Ennifar, J.E. Meyer, F. Buchholz, A.F. Stewart, D. Suck, Nucleic Acids Res. 31, 5449–5460 (2003)CrossRefGoogle Scholar
  37. 37.
    Y. Chen, U. Narendra, E.L. Iype, M.M. Cox, A.P. Rice, Mol. Cell 6, 885–897 (2000)Google Scholar
  38. 38.
    T. Biswas, H. Aihara, M. Radman-Livaja, D. Filman, A. Landy, T. Ellenberger, Nature 435, 1059–1066 (2005)CrossRefGoogle Scholar
  39. 39.
    G.D. Van Duyne, Annu. Rev. Biophys. Biomol. Struct. 30, 87–104 (2001)CrossRefGoogle Scholar
  40. 40.
    R.H. Hoess, A. Wierzbicki, K. Abremski, Nucleic Acids Res. 14, 2287–2300 (1986)CrossRefGoogle Scholar
  41. 41.
    I. Grainge, S. Pathania, A. Vologodskii, R.M. Harshey, M. Jayaram, J. Mol. Biol. 320, 515–527 (2002)CrossRefGoogle Scholar
  42. 42.
    D. Shore, J. Langowski, R.L. Baldwin, Proc. Natl. Acad. Sci. U. S. A. 78, 4833–4837 (1981)CrossRefGoogle Scholar
  43. 43.
    D.M. Crothers, J. Drak, J.D. Kahn, S.D. Levene, Methods Enzymol. 212, 3–29 (1992)Google Scholar
  44. 44.
    Q. Du, C. Smith, N. Shiffeldrim, M. Vologodskaia, A. Vologodskii, Proc. Natl. Acad. Sci. U. S. A.102, 5397–5402 (2005)CrossRefGoogle Scholar
  45. 45.
    T.E. Cloutier, J. Widom, Mol. Cell 14, 355–362 (2004)CrossRefGoogle Scholar
  46. 46.
    S.D. Levene, S.M. Giovan, A. Hanke, M.J. Shoura, Biochem. Soc. Trans. 41, 513–518 (2013)CrossRefGoogle Scholar
  47. 47.
    Y. Zhang, A.E. McEwen, D.M. Crothers, S.D. Levene, Biophys. J. 90, 1903–1912 (2006)CrossRefGoogle Scholar
  48. 48.
    R.E. Dickerson, J. Biomol. Struct. Dyn. 6, 627–634 (1989)CrossRefGoogle Scholar
  49. 49.
    M.J. Shoura, A.A. Vetcher, S.M. Giovan, F. Bardai, A. Bharadwaj, M.R. Kesinger, S.D. Levene, Nucleic Acids Res. 40, 7452–7464 (2012)CrossRefGoogle Scholar
  50. 50.
    L. Ringrose, V. Lounnas, L. Ehrlich, F. Buchholz, R. Wade, A.F. Stewart, J. Mol. Biol. 284, 363–384 (1998)CrossRefGoogle Scholar
  51. 51.
    K. Rippe, M. Guthold, P.H. von Hippel, C. Bustamante, J. Mol. Biol. 270, 125–138 (1997)CrossRefGoogle Scholar
  52. 52.
    V.A. Bloomfield, D.M. Crothers, I.J. Tinoco, Nucleic acids: structures, properties and functions (University Science Books, Herndon, 2000)Google Scholar
  53. 53.
    L. Finzi, J. Gelles, Science 267, 378–380 (1995)CrossRefGoogle Scholar
  54. 54.
    J. Muller, S. Oehler, B. Muller-Hill, J. Mol. Biol. 257, 21–29 (1996)CrossRefGoogle Scholar
  55. 55.
    T.M. Dunn, S. Hahn, S. Ogden, R.F. Schleif, Proc. Natl. Acad. Sci. U. S. A. 81, 5017–5020 (1984)CrossRefGoogle Scholar
  56. 56.
    E. de Wit, W. de Laat, Genes Dev. 26, 11–24 (2012)CrossRefGoogle Scholar

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© Springer-Verlag Berlin Heidelberg 2014

Authors and Affiliations

  1. 1.The University of Texas at DallasRichardsonUSA

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