The M center dot ADP center dot P-i state is required for helical order inthe thick filaments of skeletal muscle

The thick filaments of mammalian and avian skeletal muscle fibers are disor dered at low temperature, but become increasingly ordered into an helical s tructure 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 trans ition as reflecting a coupling between nucleotide state and global conforma tion with M . ATP (disordered) being favored at 0 degrees C and M . ADP . P -i (ordered) at 20 degrees C. However, hitherto this has been limited to a qualitative correlation and the biochemical state of the myosin heads requi red to obtain the helical array has not been unequivocally identified. In t he present study we have critically tested whether the helical arrangement of the myosin heads requires the M . ADP . P-i state. X-ray diffraction pat terns were recorded from skinned rabbit psoas muscle fiber bundles stretche d to non-overlap to avoid complications due to interaction with actin, The effect of temperature on the intensities of the myosin based layer lines an d on the phosphate burst of myosin hydrolyzing ATP in solution were examine d under closely matched conditions, The results showed that the fraction of myosin mass in the helix closely followed that of the fraction of myosin i n the M . ADP . P-i state. Similar results were found by using a series of nucleoside triphosphates, including CTP and GTP, In addition, fibers treate d 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 st ate revealed no helical order. Diffraction patterns from muscle fibers in n ucleotide-free and in ADP-containing solutions did not show helical structu re. 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 refection of the thick filament is linked to the helic al order. The spacing in the ordered M . NDP . P-i state is 143.4 Angstrom, but in the disordered state, it is 144.2 Angstrom. This may be explained b y the different interference functions for the myosin heads and the thick f ilament backbone.