Publication Date

2017-5

Document Type

Honors Thesis

Degree Name

Bachelor of Arts

Department

Biochemistry

Advisors

Nathan D. Derr

Keywords

Nanoscale cargo transport, DNA origami, Motor proteins, Single molecule microscopy, Cytoskelton, Kinesin, Dynein, Nanoelectrical systems, Microscopy, DNA, Origami

Abstract

Eukaryotic cells employ cytoskeletal motor proteins to efficiently organize and distribute intracellular cargos. To transport these cargoes, multiple motors will work together in teams; however, several questions remain regarding the biophysical mechanisms of these ensembles. The same attributes that make these motors essential tools for cells also makes them useful in bio-nanotechnology, suggesting that cytoskeletal motors could be co-opted for rationally designed molecular transport applications. DNA origami techniques provide unique tools to gain further insight into both the biophysical mechanisms and nanotechnological potential of motor proteins. For example, DNA origami can be used to make complex nanoscale shapes. Between 2009 and 2017, different groups have employed this technique to devise spherical constructs with varied curvature. Others have gone a step further to enclose cancer therapy drugs and other compounds into these structures. In this study, we reimagine these DNA origami structures for applications with molecular motors. We conjugate motor proteins to these spherical structures and explore their potential to encapsulate untethered fluorescent cargos. These spherical constructs were based upon the original DNA origami sphere of (Han et al., 2011) and (Kohman & Han, 2015) and were redesigned to include addressable fluorophore and motor protein attachment points. Formation of the spheres was confirmed via TEM microscopy and agarose gel electrophoresis. Image sequences acquired from TIRF microscopy verified their functionality and capacity to be transported by teams of kinesin and teams of dynein. Moving forward, the spherical ensembles could be used to study the biophysics of molecular motor motion and be utilized for applications in bio-nanotechnology.

Language

English

Comments

128 pages : color illustrations. Includes bibliographical references (pages 120-128)

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