DIRECT PHOSPHORYLATION OF I-KAPPA-B BY IKK-ALPHA AND IKK-BETA - DISCRIMINATION BETWEEN FREE AND NF-KAPPA-B-BOUND SUBSTRATE

A large protein complex mediates the phosphorylation of the inhibitor of kappa B (I kappa B), which results in the activation of nuclear fac tor kappa B (NF-kappa B). Two subunits of this complex, I kappa B kina se alpha (IKK alpha) and I kappa B kinase beta (IKK beta), are require d for NF-kappa B activation. Purified recombinant IKK alpha and IKK be ta expressed in insect cells were used to demonstrate that each protei n can directly phosphorylate I kappa B proteins. IKK alpha and IKK bet a were found to form both homodimers and heterodimers. Both IKK alpha and IKK beta phosphorylated I kappa B bound to NF-kappa B more efficie ntly than they phosphorylated free I kappa B. This result explains how free I kappa B can accumulate in cells in which IKK is still active a nd thus can contribute to the termination of NF-kappa B activation.