Effect of high hydrostatic pressure on chicken myosin subfragment-1

Abstract

Hydrostatic pressure-induced structural changes in subfragment-1 (S1) of myosin molecule were studied. ATP-induced emission spectra of S1 were used to detect global structural change of S1 by pressure treatment. The fluorescence intensity of unpressurized S1 increased by addition of ATP. The increment of fluorescence of pressurized S1 up to 150 MPa was almost the same as control, whereas it became smaller above 200 MPa. ATP binding ability of S1 examined using 1, N 6-ethenoadenosine 5′-diphosphate (ε-ADP) indicated that the binding of ε-ADP to S1 decreased in the range of 250–300 MPa. S1 pressurized below 250 MPa and unpressurized S1 similarly bound to F-actin, although binding of S1 pressurized above 250 MPa decreased. Electron microscopic observation revealed arrowhead structure in control acto-S1, while disordered arrowhead structure was observed in acto-S1 prepared from pressurized S1 at 300 MPa. S1 pressurized below 250 MPa retained the same actin activated ATPase activity as the control, whereas the activity decreased to 60% at 300 MPa. Pressure treated S1 was easily cleaved by tryptic digestion into three domains, i.e. 27 kDa (N-terminal), 50 and 20 kDa (C-terminal) fragments, which were the same as those in unpressurized one. It is concluded that pressure-induced global structural changes of S1 begin to occur about 150 MPa, and the local structural changes in ATPase and actin binding sites followed with elevating pressure to 250–300 MPa.