The overall goals of the proposed research are to establish the structure and mechanism of action of the dynein ATPase in generating a force for ciliary and flagellar movement. The specific goals of these studies can be divided into five parts: (1) Localize the polypeptides, the ATPase sites, and the microtubule-binding sites in terms of the three-headed bouquet of the 22S dynein; (2) evaluate the flexibility of the strands connecting the dynein heads to the base; (3) determine the arrangement of the dynein heads on the microtubule surface lattice and the nature of the interaction in the rigor complex; (4) examine the ATP-dependent changes in conformation; and (5) continue studies on the structure and localization the 14S dynein ATPase. These problems will be addressed by a combination of techniques including: transmission electron microscopic (TEM) localization of monoclonal and polyclonal antibodies; scanning transmission electron microscopy (STEM) of dynein fragments and the microtubule-dynein complex; hydrodynamic and fluorescence anisotropy measurements on isolated dynein; TEM analysis of unstained, unfixed, frozen-hydrated specimens or replicas of rotary shadowed preparations; and solution x-ray scattering studies of the microtubule-dynein complex and intact axonemes. The current work builds upon our previous results on the structure and ATPase pathway of dynein isolated from Tetrahymena cilia. These studies are expected to establish the structure of dynein in detail and to specify the structural basis for changes in conformation necessary to couple ATP hydrolysis to the interaction of the dynein crossbridge with microtubules to produce a force for ciliary movement. The work also provides a basis for analysis of dynein-like ATPases in other microtubule systems such as chromosome movement or the intracellular transport of membrane bound particles.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM032023-06
Application #
3280579
Study Section
Cellular Biology and Physiology Subcommittee 1 (CBY)
Project Start
1983-04-01
Project End
1991-03-31
Budget Start
1988-04-01
Budget End
1989-03-31
Support Year
6
Fiscal Year
1988
Total Cost
Indirect Cost
Name
Pennsylvania State University
Department
Type
Schools of Arts and Sciences
DUNS #
City
University Park
State
PA
Country
United States
Zip Code
16802
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