Figure 3.

Recovery of the NADH oxidase activity in the CCL16-B2 cells transfected with the S. cerevisiae NDI1 gene. (Top) Nontransfected CCL16-B2 cells (1 × 10⁷ cells). (Middle) NDI1-transfected CCL16-B2 cells (1 × 10⁷ cells). (Bottom) NDI1-transfected CCL16-B2 cells (3 × 10⁷ cells). Where indicated, 5 mM glutamate (Glu), 5 mM malate (Mal), 0.5 mM NADH, 5 μM rotenone (Rot), 5 mM succinate (Succ), 5 μM antimycin A (AntA), and 0.5 mM flavone were added. The cells were harvested by trypsinization and resuspended in 1 ml of a medium containing 20 mM Hepes, pH 7.1/250 mM sucrose/10 mM MgCl₂. The cells were treated with 50–150 μg of digitonin until more than 90% of the cells are stained by trypan blue. The digitonin-treated cells were washed with the same medium. Oxygen consumption was measured polarographically in 0.6 ml of the buffer containing 20 mM Hepes, pH 7.1/250 mM sucrose/10 mM MgCl₂ by using a Clark electrode in a water-jacketed chamber maintained at 37°C.