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Updated: Aug 8, 2026

Oligopeptide Competition Assay for Phosphorylation Site Determination
Published on: May 18, 2017
Direct phosphorylation of IkappaB by IKKalpha and IKKbeta: discrimination between free and NF-kappaB-bound substrate
1Laboratory of Gene Regulation and Signal Transduction, Department of Pharmacology, University of California at San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA.
Abstract:
A large protein complex mediates the phosphorylation of the inhibitor of kappaB (IkappaB), which results in the activation of nuclear factor kappaB (NF-kappaB). Two subunits of this complex, IkappaB kinase alpha (IKKalpha) and IkappaB kinase beta (IKKbeta), are required for NF-kappaB activation. Purified recombinant IKKalpha and IKKbeta expressed in insect cells were used to demonstrate that each protein can directly phosphorylate IkappaB proteins. IKKalpha and IKKbeta were found to form both homodimers and heterodimers. Both IKKalpha and IKKbeta phosphorylated IkappaB bound to NF-kappaB more efficiently than they phosphorylated free IkappaB. This result explains how free IkappaB can accumulate in cells in which IKK is still active and thus can contribute to the termination of NF-kappaB activation.
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