Cross-bridge orientation in skeletal muscle measured by linear dichroism of an extrinsic chromophore |
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Authors: | Julian Borejdo Olga Assulin Toshio Ando Susan Putnam |
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Institution: | 1. Polymer Department, Weizmann Institute of Science Rehovot, 76100, Israel;2. Cardiovascular Research Institute, University of California San Francisco, CA 94143, U.S.A. |
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Abstract: | Linear dichroism of chromophoric labels attached to myosin heads has been used to establish cross-bridge orientation in myofibrils and muscle fibers. Generalized expressions were obtained for the dichroic ratio of a circularly symmetrical assembly of chromophores viewed through high apertures. The theoretical expressions were used to estimate the angle Θ of the absorption dipole of the dye relative to the myofibrillar axis. Myosin subfragment-1 has been labeled with tetramethyl rhodamine and diffused into the I-band of myofibrils; endogenous muscle myosin has been labeled directly. Dichroism has been measured from these preparations in the absence (rigor) and presence of MgATP and its analogs. In rigor, angle Θ was 80 °. Relaxed and contracted preparations displayed no dichroism, suggesting a high degree of cross-bridge disorder. MgAMP-PNP2 and MgPPi imposed on the cross-bridges a distribution intermediate between rigor and relaxation. In the presence of MgADP the preparations showed strong dichroism of the opposite direction to that present in rigor. No detachment of the cross-bridges occurred under these conditions and the effect was not due to the rotational displacement of the attached dye by the nucleotide. It is concluded that the formation of a ternary complex myosin-MgADP-actin makes it possible to detect a large local deformation imposed on the cross-bridge by nucleotide binding, which results in a change of the spatial attitude of the mobile region of the protein by about 40 °. |
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