Fig. 3.

M80A cyt c does not efficiently activate caspases or induce the release of SMAC-DIABLO from mitochondria. (A) Decreased activation of caspase-3 by M80A. Equal concentrations of recombinant M80A or WT cyt c were added to cytosolic extracts of Jurkat cells in the presence of dATP and ATP. Caspase-3 activity was measured using the chromogenic substrate AcDEVDpNA. To verify the occurrence of competitive inhibition, M80A and WT cyt c were also assayed in combination, in several molar ratios. (B) M80A cyt c expression does not induce the release of SMAC/DIABLO from mitochondria. HeLa cells expressing M80A or WT cyt c or untransfected HeLa cells (Control) were separated into mitochondrial and cytoplasmic fractions and the levels of cyt c-GFP (Cyt c-GFP), SMAC/DIABLO and endogenous cyt c (Endog Cyt c) in each fraction were determined by WB analysis.