The applicant and his colleagues showed earlier that a myosin head (S1) can attach either to one or to two actin protomers in an actin filament, depending of whether filament is saturated or unsaturated with S1's. The objective of the project is to test a hypothesis, which was stimulated by this finding, that a major part of a power stroke of skeletal muscle consists of orientational change associated with transition of myosin head from a state in which it binds one actin to a state in which it binds to two actins. In this application they propose to the test this hypothesis by: (i) checking whether the kinetics of binding of S1 to unsaturated filament is consistent with 1 S1 binding to 2 actins; (ii) identifying the amino acid residues involved in making the second contact between S1 and actin; (iii) inducing point mutations in S1 to demonstrate the importance of the second actin binding site; (iv) testing whether cross-bridges in rigor muscle bind to two actins; (v) testing whether the two binding states can be seen during hydrolysis of ATP. The significance of this project is that it presents a critical hypothesis about a fundamental problem in contractility and proposes to test it experimentally. The idea of two binding states and that the transition between them is involved in force generation will be checked in skeletal and smooth muscle, but applies perhaps to the broader category of motor-driven processes. Testing this idea certainly will contribute to the understanding of the molecular changes occurring in acto-S1 during contraction. It is likely that the elucidation of the molecular mechanism of contraction of skeletal and smooth muscle will lead to better modalities in treating diseased muscle.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR040095-10
Application #
2899857
Study Section
Biophysical Chemistry Study Section (BBCB)
Program Officer
Lymn, Richard W
Project Start
1989-07-01
Project End
2002-03-31
Budget Start
1999-04-01
Budget End
2002-03-31
Support Year
10
Fiscal Year
1999
Total Cost
Indirect Cost
Name
University of North Texas
Department
Biochemistry
Type
Schools of Osteopathy
DUNS #
110091808
City
Fort Worth
State
TX
Country
United States
Zip Code
76107
Midde, K; Dumka, V; Pinto, J R et al. (2011) Myosin cross-bridges do not form precise rigor bonds in hypertrophic heart muscle carrying troponin T mutations. J Mol Cell Cardiol 51:409-18
Mettikolla, P; Calander, N; Luchowski, R et al. (2011) Cross-bridge kinetics in myofibrils containing familial hypertrophic cardiomyopathy R58Q mutation in the regulatory light chain of myosin. J Theor Biol 284:71-81
Borejdo, J; Ushakov, D S; Moreland, R et al. (2001) The power stroke causes changes in the orientation and mobility of the termini of essential light chain 1 of myosin. Biochemistry 40:3796-803
Reshetnyak, Y K; Andreev, O A; Borejdo, J et al. (2000) The identification of tryptophan residues responsible for ATP-induced increase in intrinsic fluorescence of myosin subfragment 1. J Biomol Struct Dyn 18:113-25
Andreev, O A; Borejdo, J (1999) Binding of myosin cross-bridges to thin filaments of rabbit skeletal muscle. Biochem Biophys Res Commun 258:628-31
Andreev, O A; Borejdo, J (1997) Interaction of the heavy and light chains of cardiac myosin subfragment-1 with F-actin. Circ Res 81:688-93
Borejdo, J; Burlacu, S (1994) Orientation of actin filaments during motion in in vitro motility assay. Biophys J 66:1319-27
Andreeva, A L; Andreev, O A; Borejdo, J (1993) Structure of the 265-kilodalton complex formed upon EDC cross-linking of subfragment 1 to F-actin. Biochemistry 32:13956-60
Andreev, O A; Andreeva, A L; Markin, V S et al. (1993) Two different rigor complexes of myosin subfragment 1 and actin. Biochemistry 32:12046-53
Andreev, O A; Andreeva, A L; Borejdo, J (1993) Polarization of fluorescently labeled myosin subfragment-1 fully or partially decorating muscle fibers and myofibrils. Biophys J 65:1027-38

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