An unique method for determining the permeability of the mitochondrial outer membrane

Mitochondrial function depends on the metabolite fluxes through the mitochondrial outer membrane. To directly measure the permeability of the outer membrane by using traditional methods is difficult due to the small volume of the intermembrane space and high permeability of the outer membrane. A method was developed to measure the permeability of the mitochondrial outer membrane by taking advantage of mitochondrial processes that consume the substrate being examined. A steady state concentration of substrate forms in the intermembrane space when the flux through the outer membrane equals the rate of substrate consumption. Thus, the rate of consumption of substrate depends on both processes and the extent of the contribution of each was sorted out by fitting to a steady-state theory. This report focuses on mitochondrial ATP phosphorylation and the determination of the permeability of the outer membrane to ADP. This approach also yields the intermembrane space [ADP] and shows how it differs from the extramitochondrial concentration.