(A) IDUP
in cell lysates expressing a SNAP-tagged target protein.
DNA was rapidly amplified in samples corresponding to known interactions:
(B) SNAP/CAII+(GL)CBS (ΔCT = 5–6), (C) SNAP/Bcl-xL+Bad or Bak (ΔCT = 8–9), (F) SNAP-FRB+rapamycin
(ΔCT = 6), and (G) FKBP-SNAP+rapamycin
(ΔCT = 6). (D) A sequence corresponding
to CAII+CBS was enriched ∼100-fold in a sample expressing
either SNAP-CAII (lane 7) or CAII-SNAP (lane 8). (E) In samples expressing
SNAP-Bcl-xL, a sequence corresponding to the interaction between Bcl-xL+Bak
was enriched ∼100-fold (lane 11), but no enrichment was observed
for a sequence corresponding to BclxL+BakL78A, a weakly binding
mutant of the Bak peptide (lane 9). (H) Overexpression of FKBP with
SNAP-FRB increased the enrichment of a sequence encoding FRB+rapamycin
by 10-fold compared to a sample transfected with SNAP-FRB alone. (I)
Overexpression of FRB with FKBP-SNAP also increased the enrichment
of a sequence corresponding to FKBP+rapamycin by 10-fold compared
to a sample transfected with FKBP-SNAP alone.