Figure 3.

hTom20-mediated insertion of VDAC into synthetic lipid membranes stimulates efflux of ATP. Sucrose-loaded LUVs with or without covalently attached hTom20 and containing 1.8 mM [³²P]ATP (20 Ci/mol) were incubated in a standard protein import reaction (Goping et al., 1998) with VDAC or pOCT transcription-translation products in the presence or absence of 1 mM NADH. At the indicated times, 20 vol of import reaction medium was added, the LUVs were collected by centrifugation, and radioactivity in the supernatant was determined as in Fig. 2 B. Shown are the average of the normalized results of three determinations with standard deviations (maximum radioactivity = 2,100 cpm). The range of total encapsulated ATP released from LUVs-Tom20 by VDAC at 15 min was 5–10%.