[4] Lipid enrichment and fusion of mitochondrial inner membranes

This chapter discusses the details of methods developed in the laboratory over the past few years which permit a significant incorporation or bulk enrichment of the mitochondrial inner membrane with a variety of membrane lipids. Related techniques, which permit the conversion of the typically small mitochondrial inner membranes to ultralarge inner membranes through a fusion process, are also reviewed. One method provides osmotically active, ultralarge (>200/μm) individual, spherical inner membranes. The lipid-enrichment procedure gives enriched inner membranes in bulk suspension whereas the fusion procedures are best suited for microanalysis of individual, enlarged inner membranes on microscope slides. Lipid-enriched inner membranes and fused inner membranes have been used in studies on the nature of the interaction of oxidation-reduction components utilizing routine reaction-kinetic approaches, fluorescence recovery after photobleaching, and flash-induced absorption anisotropy. Such studies reveal that the major oxidation-reduction components of the inner membrane diffuse laterally and independently in the membrane plane and rotate on an axis normal to the membrane plane. Lipid-enrichment and fusion methods are applied successfully on other membrane systems, such as microsomes and red blood cell ghosts and it is believed that most membrane systems can be treated similarly.