The NF-κB essential modulator (NEMO) is the master regulator of NF-κB signaling， controlling the immune and nervous systems. NEMO affects the activity of IκB kinase-β (IKKβ)， which relieves the inhibition of the NF-κB transcriptional regulation machinery. Despite major effort， there is only a very sparse， phenomenological understanding of how NEMO regulates IKKβ and shows specificity in its large range of molecular interactions. We explore the key molecular interactions of NEMO using a molecular biophysics approach， incorporating rapid-mixing stopped-flow， high-pressure， and CD spectroscopies. Our study demonstrates that NEMO has a significant degree of native structural disorder and that m... More
The NF-κB essential modulator (NEMO) is the master regulator of NF-κB signaling， controlling the immune and nervous systems. NEMO affects the activity of IκB kinase-β (IKKβ)， which relieves the inhibition of the NF-κB transcriptional regulation machinery. Despite major effort， there is only a very sparse， phenomenological understanding of how NEMO regulates IKKβ and shows specificity in its large range of molecular interactions. We explore the key molecular interactions of NEMO using a molecular biophysics approach， incorporating rapid-mixing stopped-flow， high-pressure， and CD spectroscopies. Our study demonstrates that NEMO has a significant degree of native structural disorder and that molecular flexibility and ligand-induced conformational change are at the heart of the molecular interactions of NEMO. We found that long chain length， unanchored， linear polyubiquitin drives NEMO activity， enhancing the affinity of NEMO for IKKβ and the kinase substrate IκBα and promoting membrane association. We present evidence that unanchored polyubiquitin achieves this regulation by inducing NEMO conformational change by an allosteric mechanism. We combine our quantitative findings to give a detailed molecular mechanistic model for the activity of NEMO， providing insight into the molecular mechanism of NEMO activity with broad implications for the biological role of free polyubiquitin.
