CAREER: Integrated Biosynthetic and Single-Molecule Approaches to Investigate Collective Motor Protein Dynamics

William Marsh Rice University, Houston TX

Investigators

Abstract

Collective interactions between motor proteins play an essential role in the transport of intracellular cargo by providing a means to optimize transport for the heterogeneous environment of the cytoplasm. As a result, collective biomotor transport is essential for the maintenance of healthy cellular function. However, significant challenges remain to fundamentally understand how motor proteins function collectively. These challenges largely stem from experimental difficulties in connecting structural information that describes the supramolecular properties of interacting groups of motor proteins to collective modes of intracellular transport. This research will bridge these gaps by establishing techniques that enable collective motor transport to be investigated from both a structural and dynamical standpoint. This goal will be accomplished by developing biosynthetic technologies that can be used to construct experimental model systems of interacting motor proteins. Based on self-assembled DNA nanostructures and engineered artificial proteins, these technologies will be harnessed to build molecular platforms that enable the position, number, and type of motors contained in an assembly, as well as the elasticity of motor interconnects to be encoded at the molecular level. The supramolecular architecture of the assemblies will be characterized using novel single-molecule imaging methods based on total-internal reflection microscopy (TIRFM). In addition, collective motor dynamics will be investigated using optical trapping instrumentation. This apparatus will allow the intricate stepping mechanics of interacting motor proteins to be investigated with single-molecule resolution and in real time. Together, both techniques will facilitate development of detailed structure-activity relationships that define how critical transport properties (velocities, load dependence, step size, dwell times, etc...) depend on the molecular architecture of assemblies. Because the mechano-chemistry of biomotors is known to be strongly dependent on strain, and hence on the mechanical coupling between motors, establishing these relationships will provide insight into how sharing force between motors influences their mechano-chemistry. Furthermore, by developing a platform technology to construct and probe a variety of architecturally rich systems of motor proteins, this research will allow mechanisms of collective transport that are typically masked by the complexities of cellular environments to be precisely determined. The philosophical framework of this research is also harnessed in an education plan that strives to enhance the scientific literacy of students and educators by empowering them with the knowledge base and skills necessary to confront frontier challenges in the biosciences and bioengineering. These goals will be accomplished by using this research as a foundation for creating research and education opportunities for both graduate and undergraduate students. Furthermore, this work incorporates outreach activities designed to expose high school students to college-level research and education. These activities will also serve to recruit under-represented minority groups to bioscience research and will be used to disseminate new instructional tools to high school educators.