Differential contribution of the conserved tyrosine residues to activity and structural stability of Ophiophagus hannah alpha-neurotoxins

Two cc-neurotoxins, Oh-4 and Oh-7, from king cobra (Ophiophagus hannah) Ven om were subjected to Tyr modification with tetranitromethane. Selective nit ration of Tyr(4) in Oh-4 caused a slight decrease in lethal toxicity of 11% and a decrease in nicotinic acetylcholine receptor (nAchR)-binding activit y of 28%, whereas nitration of Tyr(4) in Oh-7 resulted in an approximate to 60% decrease in lethality and nAchR-binding activity. When the Tyr(23) in Oh-4 or Tyr(22) in Oh-7 appears to be 'buried' in the toxin following furth er modification, the toxins lost their biological activity and conformation al change concurrently. Nevertheless, the dinitrated Oh-4 retained a beta-s heet structure as revealed by Co spectra and exhibited a precipitin reactio n with anti-Oh-4 sera. These results indicate that both Tyr(4) and Tyr(22) play a crucial role in the neurotoxicity of Oh-7, whereas intact Tyr(23) is involved in the manifestation of the toxicity of Oh-4 to a greater extent. In contrast to Oh-4, the conformational stability of Oh-7 depends heavily upon the integrity of Tyr(22).