Calorimetric characterization of the stable complex of myosin subfragment 1 with ADP and beryllium fluoride

Abstract

The thermal unfolding of the myosin subfragment 1 (Sl) in its stable complex with ADP and beryllium fluoride (Sl . ADP . BeF;) was studied by differential scanning calorimetry. It has been shown that the structure of the Sl molecule in the Sl ADP . BeF; complex is similar to that of Sl in its complex with ADP and orthovanadate (Sl ADP . Vi) but differs radically from that of nucleotide-free Sl and Sl in the Sl . ADP complex. It is concluded that the Sl . ADP . BeF; complex can be considered, like the Sl . ADP . Vi complex, a stable structural analogue of the myosin head ADP Pi transition state of the myosin-catalyzed ATP hydrolysis.

Differential scanning calorimetric study of the complexes of modified myosin subfragment 1 with ADP and vanadate or beryllium fluoride

“…Thus, the effects of the SH1 modification on the conformational changes of $1 induced by the formation of the S1.ADP.Vi complex depend on the sulfhydryl reagent employed; NEM is the most effective, especially in comparison with 1,5-IAEDANS or IAA. The formation of the stable S1.ADP.BeFx complex causes an effect similar to that observed for the S1.ADP.Vi complex although slightly less pronounced (Bobkov et al, 1993) (Fig. 2B).…”

“…We showed that formation of the S1.ADP-Vi complex causes a global change of $1 conformation which is reflected in pronounced increase of $1 thermal stability, in considerable increase in cooperativity of the thermal transition, and in significant change of $1 domain structure (Levitsky et al, 1992). Similar but slightly less pronounced effect was found for the formation of the S1.ADP-BeF~ complex (Bobkov et al, 1993). Thus, the studies of the $1 thermal denaturation by DSC allow to probe the conformational changes of the whole $1 molecule caused by formation of the S1.ADP.Vi and S1-ADP.BeFx complexes.…”

“…Recently we have studied the thermal unfolding of $1 in the S1.ADP.Vi and S1.ADP-BeF× complexes by means of differential scanning calorimetry (DSC) (Levitsky et al, 1992;Bobkov et al, 1993). We showed that formation of the S1.ADP-Vi complex causes a global change of $1 conformation which is reflected in pronounced increase of $1 thermal stability, in considerable increase in cooperativity of the thermal transition, and in significant change of $1 domain structure (Levitsky et al, 1992).…”

Interaction of myosin subfragment 1 with F‐actin studied by differential scanning calorimetry

“…In the presence of ADP these analogues also bind to S1 and induce a significant increase of the S1 thermal stability [8,12]. However, interaction of S 1 with F-actin leads to rapid release of BeFx or A1F4" fzom active site of the S1 ATPase [15].…”

“…S 1 was obtained by digestion of myosin filaments t~om rabbit skeletal muscles by chymotrypsin [1t]: Concentration of S1 was determined spectrophotometrically by using E TM = 7.5 at 280 urn. The ternary complexes S1-ADP-BeFx and S1-ADP-A1F4" were obtained by the methods described earlier [8,12].…”