Determining the atomic level mechanism by which muscle proteins generate force and motion remains one of the fundamental questions in physiology. The recent determinations of the x-ray structures of different conformations of the motor protein, myosin, have provided new hypotheses as to its mechanism of function. However, a unifying framework defining the interaction of the nucleotide, ATP, with myosin to generate force and motion remains undetermined. Nucleotide (ATP) analogs allow one to perturb the interaction of the substrate with the protein, and correlate the perturbations with modulation of contractile activity. This can serve as a probe of the mechanism by which nucleotide hydrolysis drives the conformational changes in myosin that result in motility. We will use molecular dynamics (MD) simulations to investigate the interaction of ATP and nucleotide analogs with myosin x-ray structures as a probe of the mechanism by which nucleotide hydrolysis drives the conformational changes in myosin that generate force and motion. Our fundamental working hypothesis is that quantitative analyses of the x-ray structures can yield insights into function that other approaches have failed to discern. The simulation of nucleotide analogs at the active site will perturb the nucleotide-protein patterns normally associated with ATP binding, and allow further structure- function correlations to be drawn regarding the interaction of protein and substrate. A major goal will be to relate the simulation analyses to existing experimental data, and to identify further experimental challenges to the hypotheses generated by our modeling studies. Additional MD simulations will investigate the thermodynamic stability of the myosin dimerization domain and other alpha-helical coiled coil structures. The stability of the dimerization domain and the implications for myosin head-head interactions and the regulation of function remain unresolved, with some models suggesting a melting of the coiled coil as crucial to function. Quantitative MD simulations will be employed to analyze the relative stabilities of the coiled-coil domains of different myosin isoforms and their relationship to function. In addition to myosin II, we will investigate myosin V and myosin VI. In the latter isoform, the presence of an unstable coiled-coli domain has been hypothesized to be crucial for function. The protocols will be extended to study mutations in the coiled-coil regulatory protein, tropomyosin, which are associated with familial hypertrophic cardiomyopathy (FHC). An atomic level understanding of muscle protein function can be expected to lead to more rational therapies for FHC and other muscle diseases. Improved understanding of the function of myosin V and myosin VI can be anticipated to improve therapies for Griscelli syndrome and myosin Vl-based sensorineural hearing loss in humans. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR053720-02
Application #
7431791
Study Section
Special Emphasis Panel (ZRG1-MOSS-K (07))
Program Officer
Nuckolls, Glen H
Project Start
2007-09-01
Project End
2011-08-31
Budget Start
2008-09-01
Budget End
2009-08-31
Support Year
2
Fiscal Year
2008
Total Cost
$251,997
Indirect Cost
Name
Washington State University
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
041485301
City
Pullman
State
WA
Country
United States
Zip Code
99164
Racki, Lisa R; Naber, Nariman; Pate, Ed et al. (2014) The histone H4 tail regulates the conformation of the ATP-binding pocket in the SNF2h chromatin remodeling enzyme. J Mol Biol 426:2034-44
Canzio, Daniele; Liao, Maofu; Naber, Nariman et al. (2013) A conformational switch in HP1 releases auto-inhibition to drive heterochromatin assembly. Nature 496:377-81
Eldred, Catherine C; Naber, Nariman; Pate, Edward et al. (2013) Conformational changes at the nucleotide site in the presence of bound ADP do not set the velocity of fast Drosophila myosins. J Muscle Res Cell Motil 34:35-42
Naber, Nariman; Cooke, Roger; Pate, Edward (2011) Slow myosin ATP turnover in the super-relaxed state in tarantula muscle. J Mol Biol 411:943-50
Harrington, Timothy D; Naber, Nariman; Larson, Adam G et al. (2011) Analysis of the interaction of the Eg5 Loop5 with the nucleotide site. J Theor Biol 289:107-15
Purcell, Thomas J; Naber, Nariman; Sutton, Shirley et al. (2011) EPR spectra and molecular dynamics agree that the nucleotide pocket of myosin V is closed and that it opens on binding actin. J Mol Biol 411:16-26
Hyatt, David; Cooke, Roger; Pate, Edward (2009) Analysis of the interaction of the nucleotide base with myosin and the effect on substrate efficacy. Biophys J 97:1952-60