We analyzed 45 batches of
venom from 20 different species belonging to 11 genera from the 3 main families of venomous snakes (Elapidae, Viperidae and Crotalidae). We found high
acetylcholinesterase (AChE) activity in all
venoms from Elapidae, except in those from the Dendroaspis genus. AChE was particularly abundant in Bungarus
venoms which contain up to 8 mg of
enzyme per gram of dried
venom. We could not detect
acetylcholinesterase activity in any batch of
venom from Viperidae or Crotalidae. Titration of active sites with an organophosphorous agent (MPT) revealed that the AChE of all
venoms have similar turnovers (6000 to 8000 s(-1)) which are clearly higher than those of Torpedo and mammalian
enzymes but lower than that of Electrophorus.
AChEs from the
venom of elapid snakes of the Bungarus, Naja, Ophiophagus and Haemacatus genera were purified by affinity chromatography. SDS-PAGE analysis and
sucrose gradient centrifugation demonstrated that AChE is exclusively present as a nonamphiphilic monomer. These
enzymes are true
AChEs, hydrolyzing
acetylthiocholine faster than
propionylthiocholine and
butyrylthiocholine and exhibiting excess substrate inhibition. Twenty-seven different
monoclonal antibodies directed against AChE from Bungarus fasciatus
venom were raised in mice. Half of them recognized exclusively the Bungarus
enzyme while the others cross-reacted with
AChEs from other
venoms. Polyspecific mAbs were used to demonstrate that
venoms from Dendroaspis, which contain the AChE inhibitor
fasciculin but lack AChE activity, were also devoid of immunoreactive AChE
protein. AChE inhibitors acting at the active site (
edrophonium,
tacrine) and at the peripheral site (
propidium,
fasciculin), as well as bis-quaternary
ligands (
BW284C51,
decamethonium), were tested against the
venom AChEs from 11 different species. All
enzymes had a very similar pattern of reactivity with regard to the different inhibitors, with the exception of
fasciculin.
AChEs from Naja and Haemacatus
venoms were relatively insensitive to
fasciculin inhibition (IC50 >> 10(-6) M), while Bungarus (IC50 approximately 10(-8) M) and especially Ophiophagus (IC50 < 10(-10) M)
AChEs were inhibited very efficiently. Ophiophagus and Bungarus
AChEs were also efficiently inhibited by a
monoclonal antibody (Elec-410) previously described as a specific
ligand for the Electrophorus electricus peripheral site. Taken together, these results show that the
venoms of most Elapidae snakes contain large amounts of a highly active non-amphiphilic monomeric AChE. All
snake venom AChEs show strong immunological similarities and possess very similar enzymatic properties. However, they present quite different sensitivity to peripheral site inhibitors,
fasciculin and the
monoclonal antibody Elec-410.