The electron transfer chain is organized into inner mitochondrial membrane supercomplexes that promote substrate channeling and catalytic efficiency. This study shows that fatty acid oxidation enzymes physically interact with electron transfer chain supercomplexes at two points. The fatty acid oxidation trifunctional protein interacts with the NADH-binding domain of complex I of the electron transfer chain, while electron transfer flavoprotein dehydrogenase interacts with electron transfer chain complex III. These findings provide first view of an integrated molecular architecture for the major energy generating pathways in mitochondria.