A METHOD FOR ESTABLISHING THE HANDEDNESS OF BIOLOGICAL MACROMOLECULES

When biological macromolecules are imaged in the transmission electron microscope (TEM), their inherent handedness is lost because the three -dimensional (3D) structure is projected onto a two-dimensional (2D) p lane, and identical 2D projections can be made from either 3D enantiom er. Nevertheless, tilt experiments in the TEM can be used to determine handedness. These experiments have been performed successfully on neg atively stained specimens. More recently, the method was applied to un stained, frozen-hydrated specimens imaged by means of cryoelectron mic roscopy (cryoTEM) methods. Tilt experiments involve recording two micr ographs of the same particles at different tilt angles, computing enan tiomeric reconstructions from particle images in one micrograph, predi cting orientations of corresponding particles in the second micrograph , and comparing model projections with particle images in the second m icrograph. In principle, this procedure can be used to determine the h andedness of any biological macromolecule imaged by cryoTEM, provided the enantiomeric reconstructions are distinguishable. (C) 1997 Academi c Press.