Figure 1. VARP directly binds SNX27 in vitro.
(A) Coomassie Blue stained SDS-PAGE gel showing GST pulldown experiments with purified GST-SNX27 as bait and VARP-H10 as prey (top panel) and ⍺-His blot (bottom panel). GST-SNX27 is sufficient to pull down full-length VARP in vitro. (B and C) Biophysical data using the BLI Octet system demonstrate VARP-H10 directly binds SNX27 (B) or Retromer (C) with low micromolar affinity with KD values below 1 μM. VARP-H10 was loaded onto a Ni-NTA biosensor, and data were obtained for either SNX27 or Retromer (positive control) at a range of concentrations. Fitted data were plotted in GraphPad Prism, and binding kinetics were calculated using the Octet R8 analysis software package.