Electron paramagnetic resonance (EPR, ST-EPR) and differential scanning calorimetry(DSC) were used in conventional and temperature
modulated mode to study internal motions and energetics of myosin in skeletal muscle fibres in different states of the actomyosin
ATPase cycle. Psoas muscle fibres from rabbit were spin-labelled with an isothiocyanate-based probe molecule at the reactive
sulfhydryl site (Cys-707) of the catalytic domain of myosin. In the presence of nucleotides (ATP, ADP, AMP⋅PNP) and ATP or
ADP plus orthovanadate, the conventional EPR spectra showed changes in the ordering of the probe molecules in fibres. In MgADP
state a new distribution appeared; ATP plus orthovanadate increased the orientational disorder of myosin heads, a random population
of spin labels was superimposed on the ADP-like spectrum.
In the complex DSC pattern, higher transition referred to the head region of myosin. The enthalpy of the thermal unfolding
depended on the nucleotides, the conversion from a strongly attached state of myosin to actin to a weakly binding state was
accompanied with an increase of the transition temperature which was due to the change of the affinity of nucleotide binding
to myosin. This was more pronounced in TMDSC mode, indicating that the strong-binding state and rigor state differ energetically
from each other. The different transition temperatures indicated alterations in the internal microstructure of myosin head
region The monoton decreasing TMDSC heat capacities show that Cp of biological samples should not be temperature independent.