Shared Ly6/uPAR structural features and epitope mapping of CD59 and the Crotalus neutralizing factor (CNF)

Structural motifs can be conserved among proteins with low primary-sequence identity, potentially supporting distinct yet related molecular functions. Endogenous phospholipase A₂ inhibitors from snake plasma (sbPLIs) include the γ-type inhibitor from Crotalus durissus terrificus, known as Crotalus neutralizing factor (CNF), which protects the snake by selectively inhibiting the lethal and PLA₂ activities of crotoxin (CTx). CD59, a complement regulator and single-LU-domain member of the Ly6/uPAR superfamily, shares the three-finger fold with CNF, raising the possibility of structural and functional convergence. Here, CD59 linear peptide arrays produced by SPOT synthesis were probed with CB, the PLA₂ subunit of CTx, to identify CD59 regions capable of binding CB. Two reactive clusters were mapped, and derived peptides were synthesized and tested for their ability to inhibit the PLA₂ activity of CTx and CB in vitro. Four CD59-derived peptides interacted with CB on the arrays; however, none inhibited PLA₂ activity, in contrast to native CNF used as a positive control. These findings indicate that, although CD59 contains linear segments that recognize CB, this interaction is not sufficient to reproduce the PLA₂-inhibitory function of CNF, suggesting that any functional interplay between these Ly6/uPAR proteins is likely to depend on more complex structural determinants.