Step Length Measurement—Theory and Simulation for Tethered Bead Constant-Force Single Molecule Assay

Anders E. Wallin (Corresponding Author), Ari Salmi, Roman Tuma

Research output: Contribution to journalArticleScientificpeer-review


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.
Original languageEnglish
Pages (from-to)795-805
JournalBiophysical Journal
Issue number3
Publication statusPublished - 1 Aug 2007
MoE publication typeA1 Journal article-refereed


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