Extended Data Fig. 6 | Validation of SKP1/FBXL17-BTB structure
through KEAP1 mutations. a, Single mutations in KEAP1 do not inhibit the
co-translational SCFFBXL17-dependent degradation of the BTB protein. 293T
cells were transfected with either wild-type (left three lanes) or mutant HAKEAP1
(right two lanes; mutations denoted on the right), as well as M YCSKP1, FBXL17FLAG
and dominant negative CUL1 (dnCUL1), as indicated on top. KEAP1 levels were
monitored by gel electrophoresis and αHA western blotting. This experiment
was performed once. b, Single mutation of residues in KEAP1 at the interface
with FBXL17 do not inhibit co-translational binding of the BTB protein to
SCFFBXL17. 293T cells were transfected with wild-type or mutant HAKEAP1,
M YCSKP1, FBXL17FLAG, and dominant-negative CUL1 (dnCUL1). FBXL17FLAG was
affinity-purified and bound proteins were detected by gel electrophoresis
and western blotting. This experiment was performed once. c, Ala109
(red stick) in KEAP1 (blue) is positioned further from FBXL17 compared to the
corresponding A60 residue in KLHL12 (green). KEAP1 and KLHL12 BTB domains
were overlain bound to FBXL17 (KEAP1, actual structure; KLHL12, model).