An Optical Torque Wrench For Studying Kinesin Dynamics

We constructed an optical tweezers instrument capable of exerting torque and measuring the angular motions of small trapped particles, based on the rotation of linear polarization of the trapping laser beam. To change polarization, we employed an electro-optic modulator (EOM), which allows for a much simpler setup than a previous design (La Porta, A. and Wang, M.D. 2004. Optical torque wrench: angular trapping, rotation, and torque detection of quartz microparticles. Phys. Rev. Lett. 19:190801). Torque is monitored by measuring the difference between circularly left-handed and right-handed components of the transmitted beam: constant torque is implemented by feeding this angular signal back into a custom-designed electronic servo loop. The limited dynamic range of the EOM (±180°) is extended by monitoring the drive signal with a microcontroller, which triggers a switch to flip the output polarization by ±180° once a pre-set threshold is reached (within 10 μs). These features enable us to maintain constant torque over unlimited rotations at high bandwidth (∼100 kHz). In addition, we developed optically birefringent, non-spherical particles suitable for this instrument using nanofabrication techniques. The polarization-sensitive method employed by the apparatus precludes the use of Wollaston prisms to perform differential interference contrast (DIC) imaging. However, by exploiting conventional video-enhancement techniques (including background subtraction, contrast enhancement, and frame averaging), we report that individual microtubules (∼25 nm in diameter) can be visualized without DIC optics at ∼5 frames per second. Altogether, our instrument allows for the simultaneous application of force and torque to the study of macromolecules of interest. We are presently extending our previous studies (Gutierrez-Medina, B., et al. 2339-Pos. Torsional properties of kinesin. Biophys. J. 2008. 94:2339-Pos) on the torsional properties of the molecular motor kinesin to investigate the effect of torque on its stepwise motion.