Mapping of CD2AP-binding site in HVD of CHIKV nsP3. (A) Sequences of two consecutive potential SH3 domain ligands (M1, M2) in HVD of CHIKV nsP3 (wt) and positions of the mutations that were introduced into EGFP CHIKVnsP3HVD expression constructs: Δ9, Δ16, P423A, R428A, Δ16 + P423A, Δ16 + R428A; “_” designates deleted residue; (B) Flp-In T-REx cells were transfected with plasmids expressing CHIKVnsP3HVD harbouring mutations indicated above the panel; 24 h later immunoprecipitation (IP) via EGFP-capturing beads was carried out, obtained samples were analysed by immunoblot for the presence of CD2AP and EGFP CHIKVnsP3HVD.