The tropomyosin domain is flexible and disordered in reconstituted thin filaments.
We have used EPR spectroscopy to study the rotational motion and orientation of tropomyosin labeled with maleimide spin-label, in skeletal muscle fibers. Fibers depleted of intrinsic myosin, troponin, and tropomyosin were reconstituted with labeled tropomyosin. The 3-7 ns mobility of the labeled domains was only slightly (2-fold) inhibited by reconstitution into fibers. No motional changes were observed on addition of troponin, irrespective of the presence of Ca2+; however, the binding of extrinsic myosin heads increased the rate of domain motion to that observed in solution. Orientational studies demonstrate a broad angular distribution of the labeled domain of tropomyosin, with respect to the fiber axis. Troponin reduces the orientational disorder, while the binding of Ca2+ to troponin partially reverses this ordering effect. Myosin S1 has no effect on the orientational distribution of tropomyosin. Overall, the observed changes are very small, implying a loose association of the probed domain of tropomyosin with the thin filament.
Szczesna
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Institute of Molecular Biophysics, Florida State University, Tallahassee 32306-3015.
Fajer
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PG
D000818:Animals; D004578:Electron Spin Resonance Spectroscopy; D018485:Muscle Fibers, Skeletal / Q000737:chemistry*; D018482:Muscle, Skeletal / Q000737:chemistry*; D009218:Myosins / Q000737:chemistry; D011487:Protein Conformation; D011817:Rabbits; D012996:Solutions; D014335:Tropomyosin / Q000737:chemistry*; D014336:Troponin / Q000737:chemistry
D012996:Solutions; D014335:Tropomyosin; D014336:Troponin; D009218:Myosins