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A New Muscle Contractile System Composed of a Thick Filament Lattice and a Single Actin Filament

Madoka Suzuki ^*, Hideaki Fujita  and Shin'ichi Ishiwata ^*  

^* Department of Physics, School of Science and Engineering, Waseda University, Tokyo, Japan; Tohoku University Biomedical Engineering Research Organization, Miyagi, Japan; and Advanced Research Institute for Science and Engineering, and Consolidated Research Institute for Advanced Science and Medical Care, Waseda University, Tokyo, Japan

Correspondence: Address reprint requests to Shin'ichi Ishiwata, Dept. of Physics, School of Science and Engineering, Waseda University, 3-4-1 Okubo, Shinjuku-ku, Tokyo 169-8555, Japan. Tel.: 81-3-5286-3437; Fax.: 81-3-5286-3437; E-mail: ishiwata{at}waseda.jp.

To bridge the gap between the contractile system in muscle and in vitro motility assay, we have devised an A-band motility assay system. A glycerinated skeletal myofibril was treated with gelsolin to selectively remove the thin filaments and expose a single A-band. A single bead-tailed actin filament trapped by optical tweezers was made to interact with the inside or the outer surface of the A-band, and the displacement of the bead-tailed filament was measured in a physiological ionic condition by phase-contrast and fluorescence microscopy. We observed large back-and-forth displacement of the filament accompanied by a large change in developed force. Despite this large tension fluctuation, we found that the average force was proportional to the overlap inside and outside the A-band up to 150 nm and 300 nm from the end of the A-band, respectively. Consistent with the difference in the density of myosin molecules, the average force per unit length of the overlap inside the A-band (the time-averaged force/myosin head was 1 pN) was approximately twice as large as that outside. Thus, we conclude that the A-band motility assay system described here is suitable for studying force generation on a single actin filament, and its sliding movement within a regular three-dimensional thick filament lattice.

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