Effect of modifications in conformation of protein molecules on structure of mitochondrial membranes

New methods for preparing tissues for electron microscopy were worked out with the aim of limiting as far as possible conformational changes in the protein molecules. The proteins were stabilized by intermolecular cross-linking with glutaraldehyde, dehydrated by short treatment with ethylene glycol, and embedded in Vestopal W or in hydroxypropylmethacrylate. Such treatment appeared to affect the native conformation of proteins less than dehydration by means of air drying with or without previous cross-linking. The mitochondrial membrane elements showed a particulate structure with a range of dimensions of the particles which closely corresponds to that of membrane proteins as judged from their molecular weights. A rectangularly shaped particle measuring 60 × 100 Å was frequently observed in the mitochondrial membranes. Intentional denaturation of the tissue by heat or low pH resulted in a membrane structure similar to that observed in conventionally prepared tissues. The results conform with the concept that cellular membranes consist of a two-dimensional array of globular protein molecules and lipoprotein complexes. A continuous light layer of uniform thickness which characterizes the unit membrane structure and which has been interpreted as consisting of a continuous lipid bilayer could be observed only after extensive denaturation of the membrane proteins. The triple-layered unit membrane pattern of membranes, therefore, appears to be the result of extensive denaturation of membrane proteins.