State transition analysis of spontaneous branch migration of the Holliday junction by photon-based single-molecule fluorescence resonance energy transfer

- Branch migration of Holliday junction (~ 0.7 nm) was observed by smFRET measurement.
- State transition trajectories were reproduced by photon-based VB-HMM analysis.
- State transitions were resolved at upper limit of time resolution (~ 1000 photons).
- Migration kinetics was specified as single- and multi-base-pair steps with rates.
- Branch migration is suitable as a standard sample for smFRET measurement.

Branch migration of Holliday junction (HJ) DNA in solution is a spontaneous conformational change between multiple discrete states. We applied single-molecule fluorescence resonance energy transfer (smFRET) measurement to three-state branch migration. The photon-based variational Bayes-hidden Markov model (VB-HMM) method was applied to fluorescence signals to reproduce the state transition trajectories and evaluate the transition parameters, such as transition rate. The upper limit of time resolution suggested in simulation was nearly achieved for the state dynamics with relatively small FRET changes, and the distinctions in the populations of different states were successfully retrieved. We also discuss the suitability of the HJ as a standard sample for smFRET dynamics measurements and data analysis.