Kinesin is the smallest known biped motor protein that uses ATP as a fuel to walk processively along the microtubule track. Its proper function is critical for many vital tasks including intracellular cargo transport and cell division. A deeper insight into how kinesin functions is thus not only important for advancing fundamental knowledge of molecular motors, but also critical for developing novel therapeutics against diseases involving impaired intracellular transport. While past advances revealed many important aspects on its global motility characteristics, physical mechanism underling its stepping motion remains unclear. A major difficulty in studying kinesin motility or motor proteins in general, is that the molecule dynamically senses and generates force to move, which is difficult to contemplate based on static structural picture only. To investigate the dynamic aspect in atomistic detail, we take a synergistic approach between molecular dynamics simulation and single-molecule experiment. Using molecular dynamics simulations, we discovered that kinesin generates force by folding of a domain, which we named the cover-neck bundle. While the proposed mechanism is supported by our single- molecule motility experiments testing kinesin mutants designed to generate less force, the experiments led to further questions regarding energetics of the force generation as well as the role of the force-generating step in the overall kinesin mechanochemical cycle. Furthermore, our preliminary simulations identified two other crucial aspects of kinesin motility: (1) the structural pathway by which mechanical strain is transmitted through the motor head to modulate the nucleotide affinity, which is important for motor head coordination, and (2) the dynamic role of the C-terminal flexible E-hook domains of the microtubule in biasing the trajectory of a motor head, which is critical for how kinesin makes a step. These issues will be thoroughly investigated by further simulations. Mutant kinesins will be generated that specifically alter the physical mechanism found in simulations, and experimentally tested using state-of-the-art optical trap systems. Outcome of this work will provide a clearer atomistic picture of the mechanics underling kinesin motility. With our previous R21-funded project as a precursor, the proposed work will be developed via strong synergy between experiments and simulations, which will be the basis upon which a host of other motor proteins will be investigated as our long-term goal.

Public Health Relevance

Deeper understanding of kinesin motility will enable better control of its behavior and motility characteristics, which will lead to novel therapeutics that target kinesin-mediated transport. Our combined approach of computa- tional modeling of macromolecular complexes and single-molecule manipulation experiment provides a platform upon which a range of subcellular motor processes of biomedical importance will be investigated.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM087677-04
Application #
8330273
Study Section
Macromolecular Structure and Function C Study Section (MSFC)
Program Officer
Gindhart, Joseph G
Project Start
2009-09-01
Project End
2014-08-31
Budget Start
2012-09-01
Budget End
2014-08-31
Support Year
4
Fiscal Year
2012
Total Cost
$222,559
Indirect Cost
$23,182
Name
Texas Engineering Experiment Station
Department
Biomedical Engineering
Type
Schools of Engineering
DUNS #
847205572
City
College Station
State
TX
Country
United States
Zip Code
77845
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Teng, Xiaojing; Hwang, Wonmuk (2016) Elastic Energy Partitioning in DNA Deformation and Binding to Proteins. ACS Nano 10:170-80
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Sturgill, Emma G; Das, Dibyendu Kumar; Takizawa, Yoshimasa et al. (2014) Kinesin-12 Kif15 targets kinetochore fibers through an intrinsic two-step mechanism. Curr Biol 24:2307-13
Teng, Xiaojing; Hwang, Wonmuk (2014) Chain registry and load-dependent conformational dynamics of collagen. Biomacromolecules 15:3019-29
Hwang, Wonmuk; Lang, Matthew J (2013) Nucleotide-dependent control of internal strains in ring-shaped AAA+ motors. Cell Mol Bioeng 6:65-73
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Ravikumar, Krishnakumar M; Hwang, Wonmuk (2011) Role of hydration force in the self-assembly of collagens and amyloid steric zipper filaments. J Am Chem Soc 133:11766-73
Lakkaraju, Sirish Kaushik; Hwang, Wonmuk (2011) Hysteresis-based mechanism for the directed motility of the Ncd motor. Biophys J 101:1105-13
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