Heat-induced Denaturation of Myosin Total Rod

The myosin total rod, which consists of smaller segments of light meromyosin and heavy meromyosin subfragment-2 (HMM S–2), prepared by limited papain digestion of rabbit myosin, was purified by Sepharose-2B column chromatography. The purified total rod was more homogeneous than any previously reported, and the sodium dodecyl sulfate (SDS) gel electrophoretic method yielded a molecular weight of 22?23 × 10⁴ (11?11.5 × 10⁴ × 2).

Transition temperatures of this purified myosin total rod obtained from the melting profile during heating were 47.5 and 55°C. The results of ORD and CD measurements showed almost full reversibility upon cooling after thermal treatment. However, the results obtained from difference spectra and fluorescence spectra showed incomplete reversibility with hysteresis.

This ostensible dichotomy concerning the structural thermostability of the rod portion of myosin molecule may mean that although ORD and CD studies show almost full reversibility of the helix-coil transition, local irreversible conformational changes, involving aromatic amino acid residues take place. This fact suggests that the renahired α-rope of the myosin total rod can exhibit different properties than the native molecule under conditions where no discernible loss in helix content occurs.