Defocused orientation and position imaging (DOPI) of myosin V

Toprak et al. 10.1073/pnas.0507134103.

Supporting Information

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Supporting Movie 7

Fig. 7. Theoretical charge-coupled device (CCD) images for a dipole (Q = 45°,F = 180°) with different defocusing amounts are shown. All of the units are in mm. Defocusing amount is taken as positive when the sample is moved toward the objective and is taken as negative when the sample is moved away from the objective. The optimal defocusing amount is usually approximately –500 nm. Numerical aperture is 1.45; pixel size is 100 nm.

Fig. 8. Leapfrogging molecule, the same one as Fig. 4B. The trace shows the unexpected »180° azimuthal rotations and the corrected angular values for the molecule. Exposure time is 0.75 s.

Fig. 9. The histogram for the magnitudes of a changes is shown. a values are averaged for each dwelling period.

Movie 1. Defocused images of many myosin V molecules walking on actin filaments.

Movie 2. Defocused images of a single myosin V molecule walking on actin, BR photobleaches after 36 s. Look at the molecule at the center of the window.

Movie 3. Alternating focused and defocused images of a myosin V molecule; the resulting trace is shown in Fig. 3A. Look at the molecule at the center of the window.

Movie 4. Alternating focused and defocused images of a myosin V molecule; the resulting trace is shown in Fig. 3B. Look at the molecule at the center of the window.

Movie 5. Alternating focused and defocused imaging of a myosin V molecule; the resulting trace is shown in Fig. 4A. Look at the molecule at the center of the window.

Movie 6. Simulation for a dipole that is making both azimuthal and axial rotations in laboratory coordinate system. The amplitudes of these transitions are chosen to be close to the changes we observed with almost all of the molecules. It is assumed that sample is moved away from the objective for 500 nm; the probe is in water; 1.45 NA oil objective is used; and pixel size is 100 nm. The emission from the dipole and corresponding CCD image change as dipole rotates.

Movie 7. Pure defocused imaging of a myosin V molecule; the resulting trace is shown in Fig. 4B. Look at the molecule at the center of the window.

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