Humanized-Single Domain Antibodies (VH/V<sub>H</sub>H) that Bound Specifically to<em> Naja kaouthia</em> Phospholipase A2 and Neutralized the Enzymatic Activity

<em>Naja kaouthia</em> (monocled cobra) venom contains many isoforms of secreted phospholipase A2 (sPLA<sub>2</sub>). The PLA<sub>2</sub> exerts several pharmacologic and toxic effects in the snake bitten subject, dependent or independent on the enzymatic activity. <em>N. kaouthia</em> venom appeared in two protein profiles, P3 and P5, after fractionating the venom by ion exchange column chromatography. In this study, phage clones displaying humanized-camel single domain antibodies (VH/V<sub>H</sub>H) that bound specifically to the P3 and P5 were selected from a humanized-camel VH/V<sub>H</sub>H phage display library. Two phagemid transfected <em>E. coli</em> clones (P3-1 and P3-3) produced humanized-V<sub>H</sub>H, while another clone (P3-7) produced humanized-VH. At the optimal venom:antibody ratio, the VH/V<sub>H</sub>H purified from the <em>E. coli</em> homogenates neutralized PLA<sub>2</sub> enzyme activity comparable to the horse immune serum against the <em>N. kaouthia</em> holo-venom. Homology modeling and molecular docking revealed that the VH/V<sub>H</sub>H covered the areas around the PLA<sub>2</sub> catalytic groove and inserted their Complementarity Determining Regions (CDRs) into the enzymatic cleft. It is envisaged that the VH/V<sub>H</sub>H would ameliorate/abrogate the principal toxicity of the venom PLA<sub>2</sub> (membrane phospholipid catabolism leading to cellular and subcellular membrane damage which consequently causes hemolysis, hemorrhage, and dermo-/myo-necrosis), if they were used for passive immunotherapy of the cobra bitten victim. The speculation needs further investigations