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. Author manuscript; available in PMC: 2018 Jan 25.
Published in final edited form as: Metallomics. 2017 Jan 25;9(1):48–60. doi: 10.1039/c6mt00163g

Figure 10.

Figure 10

A) Fe-S cluster assembly on endogenous aconitase (Aco1); dependence on Nfs1 activity and mitochondrial amounts. Top panel: Mitochondria isolated from Gal-Nfs1 repressed cells (lanes 1 and 2), or Gal-Isu1/Δisu2 repressed cells expressing exclusively yIsu1 (lanes 3 and 4) or fIscU (lanes 5 and 6) were tested for FeS cluster-forming activity. Mitochondria (1X = 100 μg of proteins) were incubated with 35S-cysteine for 45 min, recovered, and then the soluble proteins of mitochondria were separated by native gel and viewed by autoradiography. The signal represents holo-aconitase [Aco1 (Fe-35S)]. Bottom panel: Immunoblots showing protein levels of Nfs1, Isd11, and Put2 (loading control) from these mitochondria. B) Kinetics of 4Fe4S cluster assembly on endogenous aconitase. As in (A) above, mitochondria were analyzed for time dependence of Fe-35S labeled aconitase formation at 15, 30 and 45 min.