Nanobody Loop Mimetics Enhance Son of Sevenless 1‐Catalyzed Nucleotide Exchange on RAS

RAS proteins control various intracellular signaling networks. Mutations at specific locations were shown to stabilize their active guanosine triphosphate (GTP)‐bound state, which is associated with the development of multiple cancers. An attractive approach to modulate RAS signaling is through its...

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Bibliographic Details
Published in:Angewandte Chemie International Edition 2023-06, Vol.62 (24), p.e202219095-n/a
Main Authors: Van holsbeeck, Kevin, Fischer, Baptiste, Gonzalez, Simon, Gadais, Charlène, Versées, Wim, Martins, José C., Martin, Charlotte, Wohlkönig, Alexandre, Steyaert, Jan, Ballet, Steven
Format: Article
Language:English
Subjects:
Catalysis
Complementarity-determining region 3
Exchanging
Guanine nucleotide exchange factor
Guanine Nucleotide Exchange Factors - metabolism
Intracellular signalling
Kinetics
Mimicry
Nanobodies
Nuclear Family
Nucleotides
Optimization
Peptides
Peptidomimetics
Protein-Protein Interactions
Proteins
Ras protein
Signal Transduction
Signaling
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Summary:RAS proteins control various intracellular signaling networks. Mutations at specific locations were shown to stabilize their active guanosine triphosphate (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 (Nb14), 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 Nb14 with an EC50 value of 29 μM. Altogether, this study demonstrated that peptides able to modulate a protein‐protein interaction can be obtained by structural mimicry of a Nb paratope. Peptides based on the interacting loop of a nanobody were designed as downsized mimetics, by considering interactions with the RAS : son of sevenless 1 (SOS1) protein complex observed in their co‐crystal structure. A biochemical assay, flanked by X‐ray co‐crystal structures and kinetic studies, demonstrated the modulation of nucleotide exchange activity of the RAS : SOS1 complex, together with high structural mimicry of the nanobody paratope.
ISSN:1433-7851
1521-3773
DOI:10.1002/anie.202219095