UC Santa Barbara
Optical trapping and single molecule measurement technologies for the investigation of biomolecular forces and motion
- Author(s): Wilcox, Jamianne
- Advisor(s): Valentine, Megan T.
- et al.
Intracellular transport along microtubules is a process critical to cell health, and
one whose breakdown is associated with neurodegenerative disorders. In this thesis, we
describe new technologies that will enable and improve investigations of multiple types
of intracellular transport on microtubules.
Although the single molecule properties of many motor proteins have been well characterized,
their behavior when transporting biological cargoes as a group of motors is still
not well understood due to measurement challenges, as well as a lack of a model system
for systematically studying collective motor behavior in vitro. In this work, we discuss
the construction of an optical trapping setup capable of applying and measuring forces
with 2 pN accuracy. We report our design of a biomimetic droplet system that reproduces
the relevant surface properties of biological cargoes while allowing the droplets to
be used as optical trapping probes. We also present a new optical trapping calibration
technique that allows experimenters to utilize the nonlinear range of the trapping force
profile, and thereby measure the high forces developed by groups of motor proteins.
Finally, we investigate the distributions of diffusion coeffcients found for Microtubule
Associated Proteins diffusing on microtubules through Monte Carlo simulations, and
examine the subtleties of interpreting and reporting single molecule diffusion data.