The structure of the Ca 2+ ATPase as revealed by electron microscopy and image processing of ordered arrays

Abstract

Two-dimensional ordered arrays of the membrane-bound Ca2+ ATPase, were formed over a wide range of conditions (i.e., pH, ionic strength, temperature) in the presence of vanadate, and studied by electron microscopy and image processing. These ordered tubular and spherical membrane vesicles of Ca2+ ATPase could also be formed with approximately one bound ATP and between one and two nonchelatable Ca2+ bound. The tubular arrays ranged between 1 and 10 microns in length and had an average flattened diameter of 90 nm, as observed in negatively stained preparations. The basic building blocks of these ordered arrays appear to be linear ribbons of Ca2+ ATPase dimers. Fourier analysis of electron micrographs of these flattened tubes revealed a near-rectangular lattice (lattice angle 73.3 +/- 4.6 degrees with average lattice constants of a = 6.2 +/- 0.25 nm, and b = 11.5 +/- 0.30 nm). The double-stranded ribbons (i.e., parallel to a) are inclined by 56 +/- 3.7 degrees relative to the tube axis in a right-handed sense, as determined from freeze-dried metal-shadowed specimens. Computer averaging of negatively stained arrays reveals a crystallographic dimer of stain-excluding matter. The dimensions of each monomer within this dimer are consistent with established structural parameters, leading us to believe a form of the Ca2+ ATPase, capable of binding at least one ATP and of binding Ca2+ ions, may exist as a dimer in the sarcoplasmic reticulum.