Innermembrane association of three mitochondrial beta-oxidation enzymes revealed by immunoelectron microscopic technique.

Ultrastructural localization of three mitochondrial beta-oxidation enzymes, enoyl-CoA hydratase, 3-hydroxyacyl-CoA dehydrogenase, and 3-ketoacyl-CoA thiolase in rat liver was studied by a post-embedding immunocytochemical technique. Rat liver was fixed by perfusion. Vibratome sections (100 micron thick) were embedded in Lowicryl K4M. Ultrathin sections were separately incubated with antibody to each enzyme, followed by protein A-gold complex. Gold particles representing the antigenic sites for all enzymes examined were confined exclusively to mitochondria of hepatocytes and other sinus-lining cells. Peroxisomes were consistently negative for the immunolabelling. In the mitochondria the gold particles were localized in the matrical side of inner membrane. The control experiments confirmed the specificity of the immunolabelling. The results firstly indicate that the mitochondrial beta-oxidation enzymes are present in the matrix of mitochondria and associated with the inner membrane.