We propose to continue our studies on the molecular properties and the sarcomeric organization of two extremely large, major myofibrillar proteins -- titin and nebulin -- which are found in the skeletal and cardiac muscles of a wide range of vertebrate and invertebrate species. Our studies in the past years have indicated that titin and nebulin may be components of an elastic, continuous myofilament (the """"""""third"""""""" filament) -- distinct from this and thick filaments -- within the sarcomere. In the next grant period, we propose to (a) investigate further the molecular properties of titin and nebulin emphasizing the purification of native proteins, the study of domain organization, the study of immunological properties, the visualization of morphology and the investigation of their interactions with other myofibrillar proteins; (b) continue and extend our antibody localization studies to include the use of monoclonal antibodies and electron microscopic techniques, emphasizing the detection and labeling of ultrathin filaments in the sarcomere; (c) characterize the phosphorylation and calcium binding properties of titin and nebulin, emphasizing the possible involvement of these proteins in regulatory mechanisms; and (d) perform a survey of the distribution and organization of titin and nebulin in nature, emphasizing their presence in insect flight muscle and in nonmuscle cells. We hope that the biochemical and structural studies proposed here will lead to better understanding of the structure and function of contractile machinery, the mechanical properties of striated muscle and the mechanism of muscle contraction and relaxation.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK020270-10
Application #
3226685
Study Section
Molecular Cytology Study Section (CTY)
Project Start
1977-08-01
Project End
1988-07-31
Budget Start
1986-08-01
Budget End
1987-07-31
Support Year
10
Fiscal Year
1986
Total Cost
Indirect Cost
Name
University of Texas Austin
Department
Type
Schools of Arts and Sciences
DUNS #
City
Austin
State
TX
Country
United States
Zip Code
78713
Granzier, H L; Irving, T C (1995) Passive tension in cardiac muscle: contribution of collagen, titin, microtubules, and intermediate filaments. Biophys J 68:1027-44
Root, D D; Wang, K (1994) Calmodulin-sensitive interaction of human nebulin fragments with actin and myosin. Biochemistry 33:12581-91
Chen, M J; Wang, K (1994) Conformational studies of a two-module fragment of nebulin and implications for actin association. Arch Biochem Biophys 310:310-7
Granzier, H L; Wang, K (1993) Interplay between passive tension and strong and weak binding cross-bridges in insect indirect flight muscle. A functional dissection by gelsolin-mediated thin filament removal. J Gen Physiol 101:235-70
Root, D D; Wang, K (1993) Silver-enhanced copper staining of protein blots. Anal Biochem 209:15-9
Root, D D; Wang, K (1993) Kinetic silver staining and quantification of proteins adsorbed to microtiter plates. Anal Biochem 209:354-9
Granzier, H L; Wang, K (1993) Gel electrophoresis of giant proteins: solubilization and silver-staining of titin and nebulin from single muscle fiber segments. Electrophoresis 14:56-64
Granzier, H L; Wang, K (1993) Passive tension and stiffness of vertebrate skeletal and insect flight muscles: the contribution of weak cross-bridges and elastic filaments. Biophys J 65:2141-59
Koretz, J F; Irving, T C; Wang, K (1993) Filamentous aggregates of native titin and binding of C-protein and AMP-deaminase. Arch Biochem Biophys 304:305-9
Chen, M J; Shih, C L; Wang, K (1993) Nebulin as an actin zipper. A two-module nebulin fragment promotes actin nucleation and stabilizes actin filaments. J Biol Chem 268:20327-34

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