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Characterization of the IKK Signaling Complex
Author(s) -
Ingraham Richard H.,
Kroe Rachel R.,
Brickwood Janice R.,
Catron Katrina M.
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.4.a47-b
Subject(s) - iκb kinase , coiled coil , peptide , function (biology) , microbiology and biotechnology , biology , chemistry , transcription factor , nf κb , signal transduction , computational biology , biochemistry , gene
The transcription factor NF‐κB is critical to the coordination of both innate and adaptive immune responses. The IκB kinase (IKK) complex is an important regulator of NF‐κB activity. Targets that affect NF‐κB signaling represent attractive therapeutic strategies for autoimmune diseases. Understanding assembly of the IKK complex is a necessary step in elucidating the molecular function of this complex and provides insight on methods of potential therapeutic intervention. The IKK complex is composed of three subunits: IKKα, IKKβ and NEMO (IKKγ). In this paper analytical ultracentrifugation is used to characterize the homo‐oligomerization of the IKK components: α, β and γ as well as the hetero‐oligiomerization of the components. The program MULTI‐COIL was used to identify a C‐terminal region of IKKβ which immediately precedes the NBD (NEMO binding domain) with a propensity to form a coiled‐coil. The corresponding sequence from IKKα has a lower propensity. A fluorescent chimeric peptide was made which contains the predicted IKKβ coiled‐coil and the IKKα NBD. This peptide binds to a construct containing the first 200 residues of NEMO with a Kd of 90 nM. Fluorescence polarization studies show that IKKβ binds to NEMO with a similar affinity. The affinities of IKKα as well as a synthetic NBD peptide are much lower. We conclude that a coiled‐coil interaction in conjunction with the NBD contribute to IKK‐NEMO binding.