Figure 3

Turbidity assays of FUS in the presence of importins. (A) Left panel: turbidity assays with 8 μM MBP-FUS(1–500) in the presence of buffer, 8 µM Kapβ2, 8 µM Kapβ2·PYNLS and 8 µM Kapβ2 + 8 µM RanGTP. Right panel: turbidity assays of 8 µM MBP-FUS (full length) in the presence of buffer, 8 µM Kapβ2 or Kapβ2·M9M. (B) Left panel: turbidity assays of 8 μM MBP-FUS(1–500) in the presence of buffer, 8 µM Impβ, 8 µM Impβ + 8 µM IBB, 8 µM Impα/β, and 8 µM Impβ + 8 µM RanGTP. Right panel: turbidity assays of 8 µM MBP-FUS (full length) in the presence of buffer, 8 µM Impβ, 8 µM Impβ + 8 µM IBB and control of IBB alone. For turbidity assays In (A,B), importins and other proteins were added to the MBP-FUS proteins prior to addition of the Tev protease at time = 0 min, and OD_{395 nm} of the solutions measured 60 min after addition of the Tev protease. The experiments were performed at room temperature, OD_{395 nm} normalized to measurements of MBP-FUS proteins + buffer + Tev at time = 60 min, the mean of 3 replicate experiments, ± SD are shown. (C) Turbidity assays of 8 µM of three different MBP-FUS constructs in the presence of buffer or 2–16 µM Kapβ2 at 10 °C. Kapβ2 is added prior to Tev (added at time = 0 min) and OD_{395 nm} is recorded 60 min after Tev addition. OD_{395 nm} is normalized to measurements of respective MBP-FUS construct + buffer + Tev. Mean of 3 or 4 replicate experiments, ± SD is shown.