Three dimensional microscopic surface profiles of membranes reconstructed from freeze etching electrol micrographs.

A method of three-dimensional reconstruction of the surface profile of artificial and natural membranes from freeze quenched electron micrographs is presented. The method is based on the analysis of the variation in thickness of platinum layers, deposited under an oblique angle. In essence, it is reminiscent of the method of Eratosthenes to measure the earth's radius. The thickness of etch-like protrusions of membranes could be determined to an accuracy of about 3 A. True distances on curved surfaces rather than projections of distances are obtained. The method has been applied to both model membranes and biological membranes. The essential results are: 1. Detailed information on the symmetry and the molecular structure of the crystalline phases of dimyristoyl phosphatidylcholine was obtained. The microscopic surface profile of the ripple structure observed between the pretransition and the main transition was analysed. In accordance with a previous model we found that the ripple structure is caused by the spontaneous curvature of the monolayers. The surface profiles of the ripple structure and of the low temperature biaxial phase could be clearly distinguished. 2. The sizes and shapes of lipid domains formed by both thermically and charge-induced lateral phase separation were determined. This showed that the visual inspection of electron micrographs may lead to a considerable underestimation of the domain size. Conclusions may be drawn concerning the different phases formed upon lateral phase separation. 3. As a biological example, yeast cell membranes were studied. The method allows one to distinguish between different membrane-bound proteins by measuring the width-to-height ratio of the particles. The deformation of the lipid layer in the environment of the proteins may be determined. This deformation contains information about lipid-mediated long-range interactions between membrane proteins.