Hidden Markov models (HMM) have been proposed as a method of analysis for noisy single-molecule fluorescence resonance energy transfer (SM FRET) data. However, there are practical and fundamental limits in applying HMM to SM FRET data due to the short photobleaching lifetimes of fluorophores and the limited time resolution of detection devices. The fast photobleaching fluorophores yield short SM FRET time traces, and the limited detection time resolution generates abnormal FRET values, which result in systematic underestimation of kinetic rates. In this work, a HMM algorithm is implemented to optimize one set of HMM parameters with multiple short SM FRET traces. The FRET efficiency distribution function for the HMM optimization was modified to accommodate the abnormal FRET values resulting from limited detection time resolution. Computer simulations reveal that one set of HMM parameters is optimized successfully using multiple short SM FRET traces and that the degree of the kinetic rate underestimation is reduced by using the proposed modified FRET efficiency distribution. In conclusion, it is demonstrated that HMM can be used to reproducibly analyze short SM FRET time traces.
All Science Journal Classification (ASJC) codes
- Physical and Theoretical Chemistry
- Surfaces, Coatings and Films
- Materials Chemistry