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Hsp90β (S226A/S255A) inhibits Apaf-1 oligomerization and caspase-9 recruitment. Recombinant Apaf-1 (0.4 μM) was mixed with catalytically inactive caspase-9 [C9 (C287A)] (0.8 μM) and recombinant Hsp90β (1 μM; S226E/S255E and S226A/S255A) and then incubated with dATP (1 mM) and 0.1 μM or 0.4 μM of cytochrome c. After incubation, each sample was loaded onto a Superdex 200 column, and each column fraction was analyzed for Apaf-1 and C9 (C287A) by immunoblotting.
