Chemical Modification of the Inner Mitochondrial Membrane

Abstract

Abstract 1. Exposure of mitochondria and submitochondrial particles from bovine heart to [35S]diazobenzenesulfonate yielded information on the asymmetrical distribution of membrane components. Radioactive coupling factor 1 (ATPase) was isolated from submitochondrial particles and radioactive cytochrome c from mitochondria. The specific radioactivities were comparable to those observed when the soluble proteins were exposed to [35S]diazobenzenesulfonate. The low radioactivity of coupling factor 1 isolated from mitochondria and of cytochrome c isolated from submitochondrial particles is evidence for the lack of permeation of the radioactive marker from one side of the membrane to the other. 2. Cytochrome oxidase was labeled from both sides of the membrane, but the specific radioactivity was about 6-fold lower than that of a solubilized cytochrome oxidase treated with [35S]diazobenzenesulfonate. This finding indicates a transmembranous location of cytochrome oxidase. 3. Spectral and enzymatic analyses of mitochondria and submitochondrial particles after cross-linking with polylysine show that cytochrome a is located on the same side as cytochrome c and suggest that cytochrome a3 is located on the same side as coupling factor 1. 4. Exposure of membrane-bound coupling factor 1 to [35S]-diazobenzenesulfonate resulted in an asymmetrical labeling of its subunits which was distinctly different from the labeling pattern after exposure of the soluble coupling factor.