Flexibility of F-actin in Relation to Acto-H-Meromyosin Motility in vitro.

K. Mihashi, T. Oda*, R. Shimo & Y. Shikata

Graduate School of Polymathematics, Nagoya University, Nagoya, *Matsushita International Institute

Flexibility of F-actin which being droven by surface-fixed H-meromyosin in vitro was studied in the following two aspects. (1) Angle Fluctuation of the Sliding Direction of F-actin (ref. 1). Of the time sequence of images of a sliding F-actin in the presence of ATP, the position of the leading head was traced for 30 seconds, and the sliding direction was determined every second. Then the angle (theta) between two successive directions was measured, giving a histogram of the angle (theta). The histograms were obtained for F-actins of various lengths. The dispersion of the angle fluctuation was found to decrease with the length of F-actin. Theoretical analysis of the length dependence of the angle fluctuation showed first that F-actin moves not like a rigid rod but as a semi-flexible filament, and second that the same amount of energy for rotation (in the plane) is received by F-actins of various lengths. The latter implied that energy for rotation may be fed only through a certain portion of F-actin (most probably through the leading head). (2) Sliding Velocity of F-actins in the presence of ATP and GTP (ref. 2 &3). H-meromyosin heads with GTP bound drove F-actin at a very low speed (the maximum speed = 1.6 um/sec) in 25 mM KCl, 4 mM MgCl2 (pH 7.4) where H-meromyosin heads with ATP bound drove F-actin at a very high speed (the maximum equal to 6.9 um/sec). When GTP and ATP were mixed in the driving solution, there appeared a certain range of GTP & ATP concentrations where the sliding velocity of F-actin was larger than that of ATP alone. In other words, H-meromyosin heads with GTP bound (slow motor) did not impede but accelerated F-actin which being driven by H-meromyosin with ATP bound (fast motor). For this kind of the sensitization effect of GTP, hydrolysis of GTP was essential. Analysis showed that ATP and GTP sensitized mutualy in driving F-actin. The result is difficult to be explained by a rigid rod model of F-actin.

ref. 1 Statistical analysis on the angle fluctuation of the direction of a single F-actin sliding on the surface-fixed H-meromyosin. Shikata, Y., Shikata, A., Shimo, R., Takada, H., Kato, C., Ito, M., Oda, T., and Mihashi, K. (1994) Proc. of Japan Acad., vol 70, Ser.B 117-120. ref. 2 Additivity of the sliding velocity of acto-H-meromyosin in vitro droven by ATP and GTP. Oda, T., & Mihashi, K. (1993) Proc. of Japan Acad., vol 69, Ser. B. 70-71. ref. 3 Mutual sensitization of ATP and GTP in driving F-actin on the surface-fixed H-meromyosin. Oda, T., Shikata, Y., & Mihashi, K. (1996) Biophys. Chem. vol. 61 63-72.

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