Figure 5. Polyubiquitin chains directly and non-covalently induce specific binding between NEMO and A20.
(A) GST or GST-NEMO was incubated with affinity-purified A20 in the presence or absence of K63 polyubiquitin chains. After 15 min, 10% of input was withdrawn, and the remaining mixture was incubated with Glutathione Sepharose. Input and glutathione pull-down were immunoblotted using the indicated antibodies. (B) as in (A) except that, in lanes 6 & 7, polyUb was treated with isopeptidease T (IsoT). (C) As in (A), but including GST-NEMO ΔN. (D) Model of the polyubiquitin-NEMO-A20 complex. NT: N-terminus. NUB: NEMO ubiquitin binding. ZnF: zinc finger. Ub: ubiquitin.