Treatment with digitonin e Resolution of functional complexes of the respiratory chain The outer mitochondrial membrane is first removed by treatment with the detergent digitonin fragments of inner membrane are then obtained by Osmotic rupture osmotic rupture of the mitochondria and the fragments are gently dissolved in a Inner nembrane second detergent. The resulting mixture of inner membrane proteins is resolved by Outer membrane discarded lon-exchange chromatography into FF different complexes (I through Iv) of the respiratory chain, each with its unique protein composition, and the enzyme atp synthase(sometimes called Complex v) Solubilization with detergent followed by ion-exchange chromatography The isolated Complexes i through iv catalyze transfers between donors nadh F.FI and succinate), intermediate carriers (UQ and cytochrome c), and O,, as shown. ATP synthase, which is composed of peripheral F1 and integral(Fo proteins(discussed later in this chapter ) has only aTP hydrolyzing; ATPase ), not ATP NADH UQ Suc- UQUQ Cyt c Cyt e synthesizing, activity in vitro ATP ADP+ P cinate Reactions catalyzed by isolated Resolution of functional complexes of the respiratory chain. The outer mitochondrial membrane is first removed by treatment with the detergent digitonin. Fragments of inner membrane are then obtained by osmotic rupture of the mitochondria, and the fragments are gently dissolved in a second detergent. The resulting mixture of inner membrane proteins is resolved by ion-exchange chromatography into different complexes (I through IV) of the respiratory chain, each with its unique protein composition, and the enzyme ATP synthase (sometimes called Complex V). The isolated Complexes I through IV catalyze transfers between donors (NADH and succinate), intermediate carriers (UQ and cytochrome c), and O2 , as shown. ATP synthase, which is composed of peripheral (F1 ) and integral (F0 ) proteins (discussed later in this chapter), has only ATP￾hydrolyzing ;ATPase), not ATP￾synthesizing, activity in vitro