PRESSURE EFFECTS ON SARCOPLASMIC-RETICULUM - A FOURIER-TRANSFORM INFRARED SPECTROSCOPIC STUDY

The Ca2+-ATPase of sacroplasmic reticulum is irreversibly inactivated by exposure to 1.5-2.0 kbar pressure for 30-60 min in a Ca2+-free medium; mono- or decavanadate (5 mM) or to a lesser extent Ca2+ (2-20 mM) protect against inactivation (Varga et al. (1986) J. Biol. Chem. 261, 13943-13956). The structural basis of these effects was analyzed by FTIR spectroscopy of sarcoplasmic reticulum in 2H2O medium. The inactivation of the Ca2+-ATPase at 1.5-2.0 kbar pressure in a Ca2+-free medium was accompanied by changes in the Amide II region of the spectrum (1550 cm-1), that are consistent with increased hydrogen-deuterium (H-2H) exchange, and by the enhancement of a band at 1630 cm-1 in the Amide I region, that is attributed to an increase in β sheet. The frequency of the peak of the Amide 1 band shifted from about 1648 cm-1 at atmospheric pressure to 1642 cm-1 at ≅ 12.5 kbar pressure, suggesting a decrease in α helix, and an increase in β and/or random coil structures. Upon releasing the pressure, the shift of the Amide I band was partially reversed. Vanadate (5 mM), and to a lesser extent Ca2+ (2-20 mM), protected the Ca2+-ATPase against pressure-induced changes both in the Amide I and Amide II regions of the spectrum, together with the protection of ATPase activity. These observations establish a correlation between the conformation of the Ca2+-ATPase and its sensitivity to pressure. The involvement of the ATP binding domain of the Ca2+-ATPase in the pressure-induced structural changes is suggested by the decreased polarization of fluorescence of fluorescein 5′-isothiocyanate covalently attached to the enzyme. © 1990.