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. 2017 Jun 13;11(6):e0005626. doi: 10.1371/journal.pntd.0005626

Fig 8. TbCDC34 activity is modulated by compound CC0651.

Fig 8

(A) BSF cells were incubated with different concentrations of CC0651 and growth inhibition was calculated after 48 hours of culture. (B) Cells treated or untreated were fixed and analyzed for DNA content using DAPI. Samples treated with CC0651 showed a decrease of cells with 1N1K and an increase of cells with 2N2K. (C) In vitro ubiquitination assay. Recombinant HsCDC34 (left panel) or TbCDC34 (right panel) (10 μM) was incubated with 100 nM of human E1 and 5 μM of human ubiquitin for 0, 1, 2, 3 and 4 hours at 37°C, in the presence of 300 μM of CC0651 or DMSO. Reactions were stopped in SDS sample buffer without DTT and analyzed by SDS-PAGE 4–12% gels and western blotting with anti-Ubiquitin and anti-His antibodies (D) Changes in the levels of Mono, Di, Tri or Poly-ubiquitin and TbCDC34~Ub thioester formation were quantified and compared between treatments. Values are means ± SEM of 3 individual samples. *: p≤ 0.02; **: p≤ 0.001.