Manipulation of Receptor Oligomerization as a Strategy to Inhibit Signaling by TNF Superfamily Members

Warren J, et al., 7(339):ra80, Sci Signal, 2014

This article focuses on the interaction between the receptor-ligand pair known as receptor activator of nuclear factor- κB (RANK) and its ligand RANKL. RANKL is a member of the TNF superfamily of cytokines and it stimulates osteoclast formation and bone resorption. The authors engineered a single protein by encoding all three monomers of RANKL as a single polypeptide chain. By using an unbiased forward genetic approach, they identified mutants of RANKL with a 500-fold increase in affinity for RANK. The affinity parameters of RANKL variants with RANK were determined by the surface plasmon resonance (SPR) and biolayer interferometry (BLI) techniques. An Octet RED system equipped with SSA biosensors was used to capture biotinylated GST-RANKL fusion proteins. The BLI experiments helped in the determination of apparent KD values for the binding of RANK-Fc or OPG-Fc with RANKL variants.

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