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. 2008 Feb 21;27(6):827–839. doi: 10.1038/emboj.2008.23

Figure 3.

Figure 3

Interaction between NF-YA and aggregated Htt in vitro. (A) Scheme for the constructs of GST–Nhtt18Q, GST–Nhtt42Q or GST–Nhtt62Q. Recognition site for HRV-3C protease is indicated. (B) CBB staining of each purified protein. Bands for expressed proteins are indicated by arrows. (C) Interaction between Nhtt aggregates and His–NF-YA. GST–Nhtt–18Q, GST–Nhtt42Q or GST–Nhtt62Q treated with HRV-3C protease were incubated at 37°C for 18 h. During these processes, Nhtt42Q or Nhtt62Q, but not Nhtt18Q, formed aggregates. Each protein was co-incubated with His–NF-YA protein for 2 h at 4°C, and then subjected to filter trap assay. The aggregated proteins were detected with anti-NF-YA or anti-Htt antibody. (D) Co-aggregation of NF-YA with Nhtt62Q. After treatment with HRV-3C protease, 10 μg of Nhtt18Q or Nhtt62Q protein was co-incubated with 0.25 or 0.75 μg of His–NF-YA for the indicated time. The trapped aggregated proteins were detected as in panel C.