Biophys J, November 1999, p. 2665-2676, Vol. 77, No. 5

The M·ADP·P_i State Is Required for Helical Order in the Thick Filaments of Skeletal Muscle

 ^*Laboratory of Physical Biology, National Institute of Arthritis, Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, Maryland 20892 USA;  ^#Department of Physiological Sciences, Eastern Virginia Medical School, Norfolk, Virginia 23501 USA;  ^§Argonne National Laboratory, Chicago, Illinois USA; and  ^¶Department of Clinical Veterinary Sciences, University of Bristol, Bristol, United Kingdom

The thick filaments of mammalian and avian skeletal muscle fibers are disordered at low temperature, but become increasingly ordered into an helical structure as the temperature is raised. Wray and colleagues (Schlichting, I., and J. Wray. 1986. J. Muscle Res. Cell Motil. 7:79; Wray, J., R. S. Goody, and K. Holmes. 1986. Adv. Exp. Med. Biol. 226:49-59) interpreted the transition as reflecting a coupling between nucleotide state and global conformation with M·ATP (disordered) being favored at 0°C and M·ADP·P_i (ordered) at 20°C. However, hitherto this has been limited to a qualitative correlation and the biochemical state of the myosin heads required to obtain the helical array has not been unequivocally identified. In the present study we have critically tested whether the helical arrangement of the myosin heads requires the M·ADP·P_i state. X-ray diffraction patterns were recorded from skinned rabbit psoas muscle fiber bundles stretched to non-overlap to avoid complications due to interaction with actin. The effect of temperature on the intensities of the myosin-based layer lines and on the phosphate burst of myosin hydrolyzing ATP in solution were examined under closely matched conditions. The results showed that the fraction of myosin mass in the helix closely followed that of the fraction of myosin in the M·ADP·P_i state. Similar results were found by using a series of nucleoside triphosphates, including CTP and GTP. In addition, fibers treated by N-phenylmaleimide (Barnett, V. A., A. Ehrlich, and M. Schoenberg. 1992. Biophys. J. 61:358-367) so that the myosin was exclusively in the M·ATP state revealed no helical order. Diffraction patterns from muscle fibers in nucleotide-free and in ADP-containing solutions did not show helical structure. All these confirmed that in the presence of nucleotides, the M·NDP·P_i state is required for helical order. We also found that the spacing of the third meridional reflection of the thick filament is linked to the helical order. The spacing in the ordered M·NDP·P_i state is 143.4 Å, but in the disordered state, it is 144.2 Å. This may be explained by the different interference functions for the myosin heads and the thick filament backbone.

Biophys J, November 1999, p. 2665-2676, Vol. 77, No. 5
© 1999 by the Biophysical Society   0006-3495/99/11/2665/12  $2.00

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