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. Author manuscript; available in PMC: 2021 Mar 1.
Published in final edited form as: Genet Med. 2020 Jan 17;22(5):878–888. doi: 10.1038/s41436-019-0747-z

Fig. 4. GAND-associated missense variants L180P, R414Q, C420S and C420R disrupt binding to NuRD components CHD3/4/5 or MBD2/3.

Fig. 4.

a) FLAG-tagged MBD2 or MBD3 were co-expressed with HA-GATAD2B (wild type (WT) or L180P) by in vitro transcription-translation (IVT) in a rabbit reticulocyte lysate. b) The FLAG-tagged C1-C2 domains of CHD3, CHD4 or CHD5 was co-expressed with HA-GATAD2B (WT, R414Q, C420R, C420S, and G406R) in IVTs. In all experiments, FLAG-fusion proteins were immobilized on αFLAG affinity beads and used as baits to pull down the co-expressed HA-GATAD2B. As a negative control, wild-type (WT) HA-GATAD2B was added to beads to which no FLAG fusion protein had been immobilized. In each case, 10% of inputs and 50% of elutions were loaded on an SDS-PAGE and proteins were detected by Western blot, using αHA and/or αFLAG antibodies.

# = degradation product of FLAG-MBD2.