If enough oxygen is present, then pyruvate diffuses through the two membrane layers into the mitochondrial matrix and enters the cit ric acid cycle. The enzymes of this cycle all are dissolved in the matrix except for three: succinate, pyruvate, and a-ketoglutarate dehydro genase. Succinate dehydrogenase is the only enzyme that transfers hydrogen atoms to FAD rather than NAD. The succinate dehydrogenase molecule must be embedded in the inner mitochondrial membrane alongside the respiratory cytochromes and enzymes, because its carrier molecule, FADH, is permanently bound to the enzyme and cannot diffuse from one place to another as can NADH. Pyruvate and -ketoglutarate dehydrogenase both are large, multienzyme complexes with molecular weights in the millions, and are similarly embedded in the inner mito chondrial membrane. The other citric acid cycle enzymes float freely in the matrix. NADH produced in the cycle diffuses to the inner mem brane surface, where it and FADH. are reoxidized by the respiratory chain. Oxygen is reduced to , and ADP is phosphorylated to ATP, both processes occurring at the inner membrane surface.

It is the inner membrane that isolates the mitochondrion chemically from the cell in which it sits. The outer membrane is permeable to most molecules of low molecular weight. The inner membrane will allow only water, small neutral molecules, and short-chain fatty acids to pass through. It is impermeable to cations and anions, most amino acids, sucrose and other sugars, coenzyme A and its esters with acetate and succinate, and ADP, ATP, NAD, and NADH. Some of these molecules are transported back and forth by carrier molecules, or permeases.

One such permease exchanges ADP and ATP across the inner membrane on a one-for-one basis. Other shuttle molecules can transport fatty acid-coenzyme A complexes, phosphate, hydroxide ion, citrate, isocitrate, succinate, and malate, but not oxaloacetate. This impermeability of the inner membrane to oxaloacetate is the reason for the conversion of oxaloacetate to malate and back again during gluconeogenesis.