Step length measurement--theory and simulation for tethered bead constant-force single molecule assay.
Linear molecular motors translocate along polymeric tracks using discrete steps. The step length is usually measured using constant-force single molecule experiments in which the polymer is tethered to a force-clamped microsphere. During the enzymatic cycle the motor shortens the tether contour length. Experimental conditions influence the achievable step length resolution, and ideally experiments should be conducted with high clamp-force using slow motors linked to small beads via stiff short tethers. We focus on the limitations that the polymer-track flexibility, the thermal motion of the microsphere, and the motor kinetics pose for step-length measurement in a typical optical tweezers experiment. An expression for the signal/noise ratio in a constant-force, worm-like chain tethered particle, single-molecule experiment is developed. The signal/noise ratio is related to the Fourier transform of the pairwise distance distribution, commonly used to determine step length from a time-series. Monte Carlo simulations verify the proposed theory for experimental parameter values typically encountered with molecular motors (polymerases and helicases) translocating along single- or double-stranded nucleic acids. The predictions are consistent with recent experimental results for double-stranded DNA tethers. Our results map favorable experimental conditions for observing single motor steps on various substrates but indicate that principal resolution limits are set by thermal fluctuations.
Wallin
Anders E
AE
Department of Physical Sciences, University of Helsinki, Helsinki, Finland. anders.wallin@helsinki.fi
Salmi
Ari
A
Tuma
Roman
R
D001704:Biopolymers / Q000737:chemistry; D003198:Computer Simulation; D004247:DNA / Q000737:chemistry; D007700:Kinetics; D008863:Microspheres; D008962:Models, Theoretical; D009010:Monte Carlo Method; D012313:RNA / Q000737:chemistry
D001704:Biopolymers; D012313:RNA; D004247:DNA