Structure of intramembrane particles in proteoliposomes containing yeast mitochondrial ATPase.

Oligomycin-sensitive ATPase was isolated from yeast mitochondria and incorporated into phospholipid vesicles by cholate dialysis. The structure of the membranes was examined by freeze-fracture electron microscopy. Liposomes formed without the ATPase were small with diameters in the range 20 to 40 nm and they had smooth fracture faces. When the ATPase was introduced, larger vesicles formed (up to 500 nm in diameter) which bore intramembrane particles (IMPs) on their fracture faces. The IMP frequency was dependent on the ratio of protein to lipid used in the reconstitution. After unidirectional shadowing at an angle of 45 degrees, IMPs had a mean width of 12.7 nm and a mean shadow length of 5.9 nm. After rotary shadowing, the mean IMP diameter was 12.9 nm. When the amount of protein was higher than 1 mg/10 mg phospholipid in the initial incorporation mixture, the ATPase formed small hexagonal arrays in the liposome membranes. Computer-based image processing of such arrays showed that the IMPs had a centre-to-centre spacing of 13.7 nm. Calculations showed that each IMP may correspond to an oligomer of the ATPase.