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5.2.2013
Chemistry Colloquium - Zev Bryant

"Engineering molecular motors"

Presented by Zev Bryant, Stanford University

Hosted by Ruben Gonzalez

Thursday, May 2, 2013

Meet the Speaker at 1:30pm in room 328 Havemeyer
Tea & Cookies at 4:00pm in room 328 Havemeyer
Seminar at 4:30 in room 209 Havemeyer

Remote control and high-resolution measurement for molecular motors research

Engineering molecular motors with dynamically controllable properties will allow selective perturbation of mechanical processes in vivo. We previously constructed myosin motors that respond to a chemical signal by reversing their direction of motion along the polarized actin filament1. To expand the potential applications of controllable molecular motors, we have now developed myosins that shift gears in response to blue light. Using structure-guided protein engineering, we have incorporated photoreceptor domains into the lever arms of chimeric myosins, resulting in motors that robustly speed up, slow down, or switch directions upon illumination. These genetically encoded motors should be directly deployable inside living cells. Our successful designs include constructs based on two different myosin classes, and we have implemented optical velocity control in motors that move at microns/sec speeds.
Simultaneous measurements of DNA twist and extension have been used to measure physical properties of the double helix and to characterize structural dynamics and mechanochemistry in nucleoprotein complexes2,3. However, the spatiotemporal resolution of twist measurements has been limited by the use of angular probes with large rotational drags, preventing the detection of short-lived intermediates or small angular steps. Using gold rotor bead tracking (AuRBT), we have now achieved >100X improvements in time resolution relative to previous techniques. AuRBT employs gold nanoparticles as bright low-drag rotational and extensional probes, relying on new instrumentation that combines magnetic tweezers with objective-side evanescent darkfield microscopy. We have tested our methodology with benchmark measurements of DNA physical properties, and have implemented RBT torque spectroscopy4,5 with gold probes to achieve large improvements in Brownian-limited torque resolution. Finally, we have used AuRBT to observe the structural dynamics of DNA gyrase, examining conformational transitions at previously inaccessible timescales.

1.    Chen, L., Nakamura, M., Schindler, T.D., Parker, D. & Bryant, Z. Engineering controllable bidirectional molecular motors based on myosin. Nature Nanotechnology 7, 252-6 (2012).
2.    Basu, A., Schoeffler, A.J., Berger, J.M. & Bryant, Z. ATP binding controls distinct structural transitions of Escherichia coli DNA gyrase in complex with DNA. Nature structural & molecular biology 19, 538-46, S1 (2012).
3.    Bryant, Z., Oberstrass, F.C. & Basu, A. Recent developments in single-molecule DNA mechanics. Current opinion in structural biology 22, 304-12 (2012).
4.    Oberstrass, F.C., Fernandes, L.E. & Bryant, Z. Torque measurements reveal sequence-specific cooperative transitions in supercoiled DNA. Proceedings of the National Academy of Sciences of the United States of America 109, 6106-11 (2012).
5.    Oberstrass, F.C., Fernandes, L.E., Lebel, P. & Bryant, Z. Torque spectroscopy of DNA: base-pair stability, boundary effects, backbending, and breathing dynamics. Physical Review Letters 110, 178103 (2013).