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Analysis of flexural rigidity of actin filaments propelled by surface adsorbed myosin motors.

Bengtsson E, Persson M, Månsson A - Cytoskeleton (Hoboken) (2013)

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.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Health and Life Sciences, Linnaeus University, Kalmar, Sweden.

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Related in: MedlinePlus

Cut-off angle. (A) LP values for simulated (10 µm·s−1; nf = 200).and experimental data (velocity; 7.68 ± 0.26 µm·s−1, mean ± 95% CI, nf = 148) where different cut-off values for Δθ(s) are used. Experimental data (circles) along with simulations (grey plus sign) representing one flexibility state (LPtheor = 10 µm) of the actin filament. (B) Frequency distribution of angular changes between subsequent analyzed frames (frame rate 5 s−1). Dashed vertical lines at cut-off values tested.
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fig04: Cut-off angle. (A) LP values for simulated (10 µm·s−1; nf = 200).and experimental data (velocity; 7.68 ± 0.26 µm·s−1, mean ± 95% CI, nf = 148) where different cut-off values for Δθ(s) are used. Experimental data (circles) along with simulations (grey plus sign) representing one flexibility state (LPtheor = 10 µm) of the actin filament. (B) Frequency distribution of angular changes between subsequent analyzed frames (frame rate 5 s−1). Dashed vertical lines at cut-off values tested.

Mentions: One issue that was not explicitly considered in the analysis above is the exclusion of abrupt direction changes Vikhorev et al., 2008a, i.e., instantaneous angular changes greater than a certain cut-off value. These abrupt changes in direction may occur in experiments, e.g., due to the presence of rigor heads. The abrupt changes may also be simulated but this was not done in the above analysis. However, in the analysis, we nevertheless used a cut-off value of 1.5 radians for all analyzed data. The reason for excluding the abrupt direction changes is that they are assumed not to be sampled from an angular distribution that is due to thermal fluctuations of the leading end of the filament Duke et al., 1995. Instead, we attribute them mainly to pinning of the actin filaments on rigor myosin heads in the in vitro motility assay Vikhorev et al., 2008b. However, for high vfΔt/LP ratio (Fig. 2), the choice of the cut-off value may introduce systematic overestimation of the persistence length by removing the filaments with the most curved paths from the analysis. Importantly, however, the data in Fig. 3F shows that any such effect is negligible with the cut-off value routinely used here, particularly as the error due to path truncation at large vfΔt/LP ratio (see above) would operate in the same direction as the effect of the cut-off. Nevertheless, it was of interest to examine how the introduction of the cut-off value influenced the estimate of the persistence length. To this end, we tested a number of different cut-off values, both for experimental data and simulations (Fig. 4A). It can be seen in Fig. 4 that the dependence of the experimental data on the cut-off value closely mimics the results of Monte-Carlo simulations assuming one single population of actin filaments having a persistence length of ∼10 µm. This is also consistent with the well superimposed frequency distributions of the angular change between measurements for the experimental and simulated data (Fig. 4B).


Analysis of flexural rigidity of actin filaments propelled by surface adsorbed myosin motors.

Bengtsson E, Persson M, Månsson A - Cytoskeleton (Hoboken) (2013)

Cut-off angle. (A) LP values for simulated (10 µm·s−1; nf = 200).and experimental data (velocity; 7.68 ± 0.26 µm·s−1, mean ± 95% CI, nf = 148) where different cut-off values for Δθ(s) are used. Experimental data (circles) along with simulations (grey plus sign) representing one flexibility state (LPtheor = 10 µm) of the actin filament. (B) Frequency distribution of angular changes between subsequent analyzed frames (frame rate 5 s−1). Dashed vertical lines at cut-off values tested.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4230416&req=5

fig04: Cut-off angle. (A) LP values for simulated (10 µm·s−1; nf = 200).and experimental data (velocity; 7.68 ± 0.26 µm·s−1, mean ± 95% CI, nf = 148) where different cut-off values for Δθ(s) are used. Experimental data (circles) along with simulations (grey plus sign) representing one flexibility state (LPtheor = 10 µm) of the actin filament. (B) Frequency distribution of angular changes between subsequent analyzed frames (frame rate 5 s−1). Dashed vertical lines at cut-off values tested.
Mentions: One issue that was not explicitly considered in the analysis above is the exclusion of abrupt direction changes Vikhorev et al., 2008a, i.e., instantaneous angular changes greater than a certain cut-off value. These abrupt changes in direction may occur in experiments, e.g., due to the presence of rigor heads. The abrupt changes may also be simulated but this was not done in the above analysis. However, in the analysis, we nevertheless used a cut-off value of 1.5 radians for all analyzed data. The reason for excluding the abrupt direction changes is that they are assumed not to be sampled from an angular distribution that is due to thermal fluctuations of the leading end of the filament Duke et al., 1995. Instead, we attribute them mainly to pinning of the actin filaments on rigor myosin heads in the in vitro motility assay Vikhorev et al., 2008b. However, for high vfΔt/LP ratio (Fig. 2), the choice of the cut-off value may introduce systematic overestimation of the persistence length by removing the filaments with the most curved paths from the analysis. Importantly, however, the data in Fig. 3F shows that any such effect is negligible with the cut-off value routinely used here, particularly as the error due to path truncation at large vfΔt/LP ratio (see above) would operate in the same direction as the effect of the cut-off. Nevertheless, it was of interest to examine how the introduction of the cut-off value influenced the estimate of the persistence length. To this end, we tested a number of different cut-off values, both for experimental data and simulations (Fig. 4A). It can be seen in Fig. 4 that the dependence of the experimental data on the cut-off value closely mimics the results of Monte-Carlo simulations assuming one single population of actin filaments having a persistence length of ∼10 µm. This is also consistent with the well superimposed frequency distributions of the angular change between measurements for the experimental and simulated data (Fig. 4B).

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.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Health and Life Sciences, Linnaeus University, Kalmar, Sweden.

Show MeSH
Related in: MedlinePlus