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Inhibition of NF‐κB activation with designed ankyrin‐repeat proteins targeting the ubiquitin‐binding/oligomerization domain of NEMO
Author(s) -
Wyler Emanuel,
Kaminska Monika,
Coïc YvesMarie,
Baleux Françoise,
Véron Michel,
Agou Fabrice
Publication year - 2007
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1110/ps.072924907
Subject(s) - iκb kinase , ubiquitin , ankyrin repeat , chemistry , signal transduction , microbiology and biotechnology , kinase , nf κb , biochemistry , biology , gene
The link between the NF‐κB signal transduction pathway and cancer is now well established. Inhibiting this pathway is therefore a promising approach in the treatment of certain cancers through a pro‐apoptotic effect in malignant cells. Owing to its central role in the pathway, the IκB kinase (IKK) complex is a privileged target for designing inhibitors. Previously, we showed that oligomerization of NEMO is necessary for IKK activation and defined a minimal oligomerization domain (CC2‐LZ) for NEMO, and we developed NEMO peptides inhibiting NF‐κB activation at the level of the IKK complex. To improve the low‐affinity inhibitors, we used ribosome display to select small and stable proteins with high affinity against the individual CC2‐LZ because the entire NEMO protein is poorly soluble. Several binders with affinities in the low nanomolar range were obtained. When expressed in human cells, some of the selected molecules, despite their partial degradation, inhibited TNF‐α‐mediated NF‐κB activation while having no effect on the basal activity. Controls with a naive library member or null plasmid had no effect. Furthermore, we could show that this NF‐κB inhibition occurs through a specific interaction between the binders and the endogenous NEMO, resulting in decreased IKK activation. These results indicate that in vitro selections with the NEMO subdomain alone as a target may be sufficient to lead to interesting compounds that are able to inhibit NF‐κB activation.