Structural dynamics of individual Holliday junctions

Abstract

The four-way DNA (Holliday) junction is the central intermediate of genetic recombination, but the dynamic aspects of this important structure are presently unclear. Although transitions between alternative stacking conformers have been predicted, conventional kinetic studies are precluded by the inability to synchronize the junction in a single conformer in bulk solution. Using single-molecule fluorescence methodology we have been able to detect these transitions. The sequence dependence, the influence of counterions and measured energetic barriers indicate that the conformer transition and branch migration processes share the unstacked, open structure as the common intermediate but have different rate-limiting steps. Relative rates indicate that multiple conformer transitions occur at each intermediate step of branch migration, allowing the junction to reach conformational equilibrium. This provides a mechanism whereby the sequence-dependent conformational bias could determine the extent of genetic exchange upon junction resolution.

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Figure 1: Holliday junction structure.
Figure 2: Single-molecule time records and analysis
Figure 3: Na+ and Mg2+ influence the conformer transition rates differently.
Figure 4: Energetics of Holliday junction dynamics.

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Acknowledgements

We thank I. Rasnik for experimental help. Funding was provided by the NIH, NSF, Searle Scholars Award (T.H.) and by Cancer Research UK (D.M.J.L.). S.A.M. was partially supported by the NIH molecular biophysics training grant.

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Correspondence to Taekjip Ha.

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The authors declare no competing financial interests.

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McKinney, S., Déclais, A., Lilley, D. et al. Structural dynamics of individual Holliday junctions. Nat Struct Mol Biol 10, 93–97 (2003) doi:10.1038/nsb883

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