Proteomic characterization and lethality of the venom of the Black Judean scorpion, Hottentotta judaicus (Buthidae): expanded toxin diversity and revisited toxicological significance

Abstract

The scorpion Hottentotta judaicus inhabits the Levant region of the Middle East, including Lebanon, Jordan, Palestine,and Israel. While previous research focused on its insecticidal properties and sodium-channel-targeting toxins, its venomremains largely unexplored using modern proteomic approaches. We analyzed the venom composition of H. judaicus fromLebanese specimens using nESI-MS/MS, MALDI-TOF MS, SDS-PAGE, and RP-HPLC. Venom lethality in mice wasassessed (LD₅₀ = 11.87 [6.59–17.16] mg/kg, i.p.), confirming moderate toxicity to vertebrates. RP-HPLC on C18resolved37 peaks, with 25 eluting between 20–40% acetonitrile. Reducing SDS-PAGE revealed predominant components < 10 kDaand minor bands at 31, 46, and 77 kDa. MaLDI-TOF MS detected 20 components from 1,000–12,000 m/z. A bottom-upshotgun nLC-MS/MS approach, following in-gel tryptic digestion of venom, identified 55 components across 15 proteinfamilies. Ion channel-active toxins [K⁺ (7), Na⁺ (16), Cl⁻ (1), ryanodine receptor (1)] and enzyme components (17) werepredominant. This study provides proteomic evidence of H. judaicus venom components previously only identified at thetranscriptomic level and reveals a richer venom profile than anticipated. Novel identified components include alternativeβ-subunits of lipolysis-activating proteins, as well as homologs of Olivierus martensii antimicrobial peptide inhibitor HAP-1, Leiurus hebraeus Lqhβ1, Parabuthus transvaalicus Birtoxin, and peptide Hj2a from Hottentotta jayakari exhibiting dualα/β-toxin activity on Nav1.1 channels. This expanding repertoire of potential bioactive components prompts a reevaluationof the pathophysiological consequences of H. judaicus envenomation in humans and further exploration of their potentialbiomedical applications.