The overall aims of this research are to understand the molecular mechanism by which muscle proteins convert chemical energy into mechanical work, and to obtain a precise correlation between the physiological, biochemical and structural events of muscular contraction. Novel methods will be applied to striated muscle fibers to probe the relations between biochemical reactions of the contractile proteins, the elementary mechanical steps of the cross-bridge cycle and the corresponding structural motions. Rapid changes in the chemical concentrations of pertinent biochemical species, such as ATP, will be made by laser pulse photolysis of photolabile """"""""caged"""""""" precursors. A newly developed method to secure the fiber ends will be used to improve the uniformity and reproducibility of the contractions. The orientation of fluorescent molecules covalently bound to the myosin heads will be monitored at high time resolution to determine the rates and identity of specific structural changes of the protein molecules. This method combined with laser photolysis, new chromophoric probes, and additional labelling sites made available by recombinant technology will provide orientation and mobility information from several regions of the myosin head. The experiments will be carried out on single muscle fibers of rabbit psoas and frog semitendinosus muscles. Results from this project should significantly advance knowledge of the contractile process and thus bring a greater understanding of both normal and pathological states of striated muscle and other types of cell motility.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
4R37AR026846-18
Application #
2006070
Study Section
Special Emphasis Panel (NSS)
Project Start
1980-04-01
Project End
2002-05-31
Budget Start
1997-07-15
Budget End
1998-05-31
Support Year
18
Fiscal Year
1997
Total Cost
Indirect Cost
Name
University of Pennsylvania
Department
Physiology
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Beausang, John F; Sun, Yujie; Quinlan, Margot E et al. (2012) Orientation and rotational motions of single molecules by polarized total internal reflection fluorescence microscopy (polTIRFM). Cold Spring Harb Protoc 2012:
Beausang, John F; Sun, Yujie; Quinlan, Margot E et al. (2012) Preparation of filamentous actin for polarized total internal reflection fluorescence microscopy (polTIRFM) motility assays. Cold Spring Harb Protoc 2012:
Beausang, John F; Sun, Yujie; Quinlan, Margot E et al. (2012) Construction of flow chambers for polarized total internal reflection fluorescence microscopy (polTIRFM) motility assays. Cold Spring Harb Protoc 2012:712-5
Beausang, John F; Sun, Yujie; Quinlan, Margot E et al. (2012) The polarized total internal reflection fluorescence microscopy (polTIRFM) twirling filament assay. Cold Spring Harb Protoc 2012:719-21
Beausang, John F; Sun, Yujie; Quinlan, Margot E et al. (2012) Fluorescent labeling of calmodulin with bifunctional rhodamine. Cold Spring Harb Protoc 2012:
Beausang, John F; Sun, Yujie; Quinlan, Margot E et al. (2012) Fluorescent labeling of myosin V for polarized total internal reflection fluorescence microscopy (polTIRFM) motility assays. Cold Spring Harb Protoc 2012:
Beausang, John F; Sun, Yujie; Quinlan, Margot E et al. (2012) The acquisition and analysis of polarized total internal reflection fluorescence microscopy (polTIRFM) data. Cold Spring Harb Protoc 2012:722-5
Beausang, John F; Sun, Yujie; Quinlan, Margot E et al. (2012) The polarized total internal reflection fluorescence microscopy (polTIRFM) processive motility assay for myosin V. Cold Spring Harb Protoc 2012:716-8
Arsenault, Mark E; Sun, Yujie; Bau, Haim H et al. (2009) Using electrical and optical tweezers to facilitate studies of molecular motors. Phys Chem Chem Phys 11:4834-9
Syed, Sheyum; Snyder, Gregory E; Franzini-Armstrong, Clara et al. (2006) Adaptability of myosin V studied by simultaneous detection of position and orientation. EMBO J 25:1795-803

Showing the most recent 10 out of 26 publications