Engineering applications of kinesin motor proteins and microtubule filaments

Hybrid bionanodevices utilize biological nanoscale components to provide critical functions in a synthetic environment. These devices merge the materials of biology and the techniques of nanotechnology in revolutionary combinations. Kinesin motor proteins have evolved in nature to attach to microtubule filaments, as well as to a variety of cargoes and transport them with high efficiency. These motors and filaments can be employed in microfabricated synthetic environments to develop engineering applications. Two such applications which utilize motor-filament attachment and motor powered active transport are described in this thesis.;First, a novel quantification technique which measures protein non-fouling surfaces at extremely low protein coverages is reported. The test surfaces are initially exposed to kinesin motors (as probes). Subsequently, the measurement of the landing rate of fluorescently labeled microtubules (as markers) enables the determination of kinesin density. This characterization technique lowers the detection limit of established protein density measurement techniques (currently at ∼1 ng/cm2) by a hundred fold. Ultra-low limits of detection, dynamic range, ease of detection and availability of a ready-made kinesin microtubule kit makes this technique highly suitable for detecting protein adsorption below the detection limits of standard techniques.;Secondly, utilization of molecular shuttles in the construction of smart dust devices is reported. Molecular shuttles achieve controlled transport of nanoscale cargo by patterning kinesin motors on a surface and employing functionalized microtubules, which are propelled by these motors, as cargo carrying elements. In this work, the challenge of cargo loading onto the shuttles is addressed. It is demonstrated that as a result of glue-like character of biotin-streptavidin bonds, the speed of streptavidin-coated microtubules has to be optimized to facilitate attachment of biotinylated cargo. The results from these investigations then enable the design of a functional smart dust biosensor powered exclusively by molecular shuttles. The device autonomically tags, transports, deposits and detects unlabeled analytes. Molecular shuttles capture antigens from the test solution, move until they bind fluorescent markers and accumulate at a distinct location for fluorescent read-out. The device does not need pressure-driven fluid flow or electro-osmotic flow to drive the mass transport functions -- a critical impediment in nanoscale devices.