Abstract
We have designed single-molecule FRET encoders which convert relative motion between two individual biomolecules to a periodic signal. Fundamental signal frequencies obtained with individual DnaB helicase molecules imply unwinding velocities ranging from 200 to 1100 base pairs per second, while low-frequency modulation of peak heights may suggest azimuthal rotation of the helicase. Signal durations show that a single helicase is capable of unwinding many hundreds of base pairs before dissociating. The initial scheme chosen for FRET encoder synthesis was expensive and limited to 5 periods, restricting the duration of the collected signal. We have utilized polymerization-driven self-assembly and rolling circle replication to synthesize inexpensive, extended FRET encoders as long as 150 periods.
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