Abstract
We compared thermally induced denaturation and aggregation of two isoforms of the isolated myosin head (myosin subfragment 1, S1) containing different “essential” (or “alkali”) light chains, A1 or A2. We applied differential scanning calorimetry (DSC) to investigate the domain structure of these two S1 isoforms. For this purpose, a special calorimetric approach was developed to analyze the DSC profiles of irreversibly denaturing multidomain proteins. Using this approach, we revealed two calorimetric domains in the S1 molecule, the more thermostable domain denaturing in two steps. Comparing the DSC data with temperature dependences of intrinsic fluorescence parameters and S1 ATPase inactivation, we have identified these two calorimetric domains as motor domain and regulatory domain of the myosin head, the motor domain being more thermostable. Some difference between the two S1 isoforms was only revealed by DSC in thermal denaturation of the regulatory domain. We also applied dynamic light scattering (DLS) to analyze the aggregation of S1 isoforms induced by their thermal denaturation. We have found no appreciable difference between these S1 isoforms in their aggregation properties under ionic strength conditions close to those in the muscle fiber (in the presence of 100 mM KCl). Under these conditions kinetics of this process was independent of protein concentration, and the aggregation rate was limited by irreversible denaturation of the S1 motor domain.
Highlights
Cyclic association-dissociation of myosin and actin, coupled with myosin-catalyzed ATP hydrolysis, is the most essential event in muscle contraction
Some difference between the two subfragment 1 (S1) isoforms was only revealed by differential scanning calorimetry (DSC), independently of the ionic strength conditions, in the thermal denaturation of the regulatory domain
The interaction of the N -terminal extension of the A1 light chain with the motor domain seems to be too weak to produce any effect on its thermal denaturation even at low ionic strength
Summary
Cyclic association-dissociation of myosin and actin, coupled with myosin-catalyzed ATP hydrolysis, is the most essential event in muscle contraction. That the interaction of the N -terminal portion of the A1 light chain with actin decreased markedly as the ionic strength was increased up to 120 mM [7] Another interesting property of the N -terminal extension of the A1 light chain is its interaction with the globular motor domain of the myosin head [12,13,14]. 35–80) and predicted a significant role of this binding in the actin-myosin interaction [14] Note that all these experiments were performed at relatively low ionic strength (up to 25–50 mM). The interaction of the N -terminal extension of the A1 light chain with the motor domain seems to be too weak to produce any effect on its thermal denaturation even at low ionic strength
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
