Nanobody CDR3 mimetics enhance SOS1-catalyzed nucleotide exchange on RAS

Abstract

RAS proteins control various intracellular signaling networks. Mutations at specific locations were shown to stabilize their active GTP-bound state, which is associated with the development of multiple cancers. An attractive approach to modulate RAS signaling is through its regulatory guanine nucleotide exchange factor (GEF) son of sevenless 1 (SOS1). With the recent discovery of Nanobody14, which potently enhances SOS1-catalyzed nucleotide exchange on RAS, we explored the feasibility of developing peptide mimetics by structurally mimicking the complementarity-determining region 3 (CDR3). Guided by a biochemical GEF assay and X-ray co-crystal structures, successive rounds of optimization and gradual conformational rigidification led to CDR3 mimetics showing half of the maximal activation potential of the native nanobody. Altogether, this study provides the first proof-of-concept that peptides able to functionally modulate a protein-protein interaction can be obtained by structural mimicry of a nanobody paratope.

Competitive, connective and allosteric Nanobodies that modulate the SOS1•RAS protein-protein interactions and tune the nucleotide exchange rate as a starting point for drug design

“…Accordingly, we synthesized peptidomimetics of the CDR3 of the most potent Nb14. Several rounds of optimization by rational design allowed us to generate potent peptides sharing similar in vitro and in cellulo effects compared to the original Nb, thereby providing the first proof-of-concept of Nb-based peptidomimetics able to functionally modulate a protein-protein interaction 25 .…”