Having established the specificity of MN551 for covalent modification of Cys111 of SOCS2, we turned our attention to the intra-SOCS family selectivity of MN551. We performed a multiple sequence alignment of the SH2 domains from all the SOCS proteins (Figure ). The Cys111 targeted by MN551 for covalency is part of the EF loop in SOCS2. Beyond SOCS2, SOCS4, SOCS5, SOCS7 and CISH also have a cysteine in the same loop (Figure ), and given the expected conserved ligand binding mode, they could all be engaged by MN551. To verify our hypothesis, we incubated recombinant SBC complexes of SOCS4, SOCS6 and CISH with MN551 for 2 hours and analysed for covalent modification of the SOCS proteins by intact ESI-MS (Figure ). These proteins were chosen as SOCS4 and 6 are already structurally well characterized, and because SOCS4 Cys350, and CISH Cys144 are in a similar position as Cys111 in SOCS2, whereas SOCS6 is lacking cysteines in the EF loop . We observed 18% modification of SOCS6 and complete modification of CISH. In contrast, MN551 failed to covalently engage SOCS4. An overlap of crystal structures of SOCS2 and SOCS4 show that although the Cys350 from SOCS4 is in the same loop as Cys111 (SOCS2), the MN551 binding mode and distance from the reactive group of MN551 may not be ideal for covalent bond formation (Figure ). Although SOCS6 lacks a cysteine in its EF loop, Cys471 from the BG loop is predicted to be in close proximity to the chloroacetamide functional group of MN551, as evident from the superposed SOCS2 and SOCS6 structures, potentially explaining the modification, albeit minimal (Figure ). Structure-guided improvements upon this observed MN551-SOCS6 covalency could expedite future attempts in development of covalent binders for SOCS6. In the absence of a PDB structure for CISH, we overlayed SOCS2 SH2 domain with an AlphaFold model generated for the CISH SH2 domain with deletions in the N-terminus and the PEST sequence (66-258, Δ174-202, Figure ). Amongst all the SOCSs, CISH has the highest sequence identity to SOCS2 SH2 and the presence of a conserved Cys144 in the EF loop together likely explain its complete covalent modification by MN551.