Analysis of flexural rigidity of actin filaments propelled by surface adsorbed myosin motors.
Bottom Line: Actin filaments are central components of the cytoskeleton and the contractile machinery of muscle.The filaments are known to exist in a range of conformational states presumably with different flexural rigidity and thereby different persistence lengths.Our results analyze the approaches proposed previously to measure the persistence length from the statistics of the winding paths of actin filaments that are propelled by surface-adsorbed myosin motor fragments in the in vitro motility assay.
Affiliation: Faculty of Health and Life Sciences, Linnaeus University, Kalmar, Sweden.Show MeSH
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Mentions: We now performed simulations (LPtheor = 10 µm) with different numbers of independent filament paths. This allowed us to test how many such paths are needed for accurate and reproducible determination of the persistence length, following the approach in the “Materials and Methods” section. Figure 2 shows estimates of path persistence lengths based on different numbers (in the range 50–2000) of simulated independent filament paths, each of 1 s duration (velocity 10 µm·s−1). For each number of independent paths, 10 different simulations were performed and for each data set, LP was obtained from an exponential fit to the CCF (Eq. 1). It can be seen in Fig. 2, that if more than about 200 and less than 1000 paths were followed, none of the 10 individual estimates for each given number of paths was different from LPtheor by more than 1.8 µm. When 1000–2000 independent paths were studied, only two out of 10 individual estimates differed from LPtheor by more than 0.5 µm. Data pooled over path length intervals of 1 or 0.5 μm were averaged and located in the midpoint of the interval in fitting the CCF. Virtually indistinguishable fits were obtained using the two different intervals (data not shown). We also tested whether the 95% CI for the exponential fit of the CCF could be used as indicator of the accuracy of the LP determination. It is clear from Fig. 2B that the individual 95% CIs obtained in fits based on 100 or less independent paths only rarely (most certainly not in 95% of the cases) are wide enough to include also the true LP. Therefore, the uncertainty in the estimates of LP from few independent paths is considerably more substantial (Fig. 2A) than indicated by the 95% CI.
Affiliation: Faculty of Health and Life Sciences, Linnaeus University, Kalmar, Sweden.