GST Affinity Pull-Down Assays.

Indicated proteins (28 μM each) were incubated at room temperature for 10 min in 25 μL of binding buffer consisting of 50 mM Tris⋅HCl (pH 8.0), 200 mM NaCl, and 2 mM DTT before addition of 30 μL G4B resin. After 10 min of incubation, the resin was spun down, and the ∼25 μL supernatant, marked as the unbound (U) fraction in the figures, was removed. The resin was then washed three times with 0.6 mL of binding buffer and the G4B-bound proteins, marked as the bound (B) fractions in the figures, were eluted with 35 μL buffer consisting of 50 mM Tris⋅HCl (pH 8.0), 200 mM NaCl, and 50 mM glutathione. One-third of each of the supernatant and eluted fractions was analyzed by SDS/PAGE and Coomassie staining.