In vitro assessment of RAS GAP activity
To estimate RAS GAP activity, HA-H-RAS together with wild-type NF or variant NF were expressed in COS-7 or HEK 293T cells. GTP-bound RAS was subsequently isolated using glutathione-agarose beads coated with recombinant GST-RAF-RBD [van Triest et al., 2001]. See Supplementary Information for details.