Cobra ( Naja spp. ) Nicotinic Acetylcholine Receptor Exhibits Resistance to Erabu Sea Snake ( Laticauda semifasciata ) Short-Chain a-Neurotoxin

Abstract

Snake alpha-neutotoxins of Elapidae venoms are grouped into two structural classes, short-chain and long-chain alpha-neutotoxins. While these two classes share many chemical and biological characteristics, there are also distinct dissimilarities between them, including their binding site on the nicotinic acetylcholine receptor (nAChR), specificity among species of Chordata, and the associated pharmacological effects. In the present study we test the hypothesis that structural motifs that evolved to confer natural resistance against conspecific long-chain alpha-neurotoxins in Elapidae snakes also interfere with the biological action of short-chain alpha-neurotoxins. We expressed functional nAChRs that contains segments or single residues of the Elapidae nAChR ligand binding domain and tested the effect of short-chain alpha-neurotoxin erabutoxin-a (ETX-a) from the Erabu sea snake Laticauda semifasciata on the acetylcholine-induced currents as measured by two-microelectrode voltage clamp. Our results show that the Elapidae nAChR alpha subunit segment T(154)-L(208) ligand binding domain has an inhibitory effect on the pharmacological action of ETX-a. This effect is primarily attributed to the presence of glycosylation at position N(189). If the glycosylation is removed from the T(154)-L(208) segment, the nAChR will be inhibited, however, to a lesser extent than seen in the mouse. This effect correlates with the variations in alpha-neurotoxin sensitivity of different species and, importantly, reflects the evolutionary conservation of the binding site on the nAChR polypeptide backbone per se. Phylogenetic analysis of alpha-neurotoxin resistance suggests that alpha-neurotoxin-resistant nAChR evolved first, which permitted the evolution of snake venom alpha-neurotoxins. A model describing alpha-neurotoxin resistance in Elapidae snakes is presented.

Rapid Radiations and the Race to Redundancy: An Investigation of the Evolution of Australian Elapid Snake Venoms

“…Large enzymes are hard to vary without loss of catalytic activity. Although receptors may modify their shape in order to achieve resistance to peptide toxins that would otherwise bind them [36], venoms of snakes favouring peptides often contain large isotypic arrays of these toxins, which may hinder the evolution of resistance in this fashion. This redundancy of targeting (see below) may first arise via the fixation of duplicate genes as the result of selection for increased dosage—the activity of peptide toxins is stoichiometric and thus relatively high concentrations of them are required to inflict a systemic impact on an envenomed prey animal.…”

A designer ligand specific for Kv1.3 channels from a scorpion neurotoxin-based library

“…3 we chose KTX, a scorpion venom peptide with an ␣-KTx scaffold that blocks Kv1.3 channels by a well-defined mechanism, as the lead for library design. ␣-KTx toxins bind directly in the potassium ion conduction pore to occlude the pathway (16) with affinities that are exquisitely sensitive to residues on the toxin and channel interaction surfaces (21,24,25). It follows that specific binding of phage to achieve library sorting demands that toxin variants (i) are synthesized and fold correctly (after proteolytic cleavage of the leader sequence that mediates surface expression), (ii) are accessible to target from the phage surface, and (iii) bind target in a stable manner despite their phage cargo.…”

“…As a control, phage expressing a mutant KTX (DDD-KTX) that does not bind to Kv1.3 were also produced. DDD-KTX has three negatively charged Asp residues at sites where KTX has basic residues critical for binding: Arg 24 , Arg 31 , and Lys 27 -the last a conserved residue in ␣-KTx toxins with an -amino group that penetrates the ion conduction pore (16).…”

Procoagulant Adaptation of a Blood Coagulation Prothrombinase-like Enzyme Complex in Australian Elapid Venom

“…Another mechanism that protects the snake against its own venom toxins is through structural modulation of the target molecule. One of these examples can be found in the Egyptian cobra ( N. haje ) that has a unique N-glycosylation site in the ligand binding domain of the nicotinic acetylcholine receptor, which has been demonstrated to obstruct binding by a venom neurotoxin, but allows binding by its natural ligand [52]. We can only speculate as to whether elapid prothrombin has undergone a similar structural modification, since its sequence is not yet available.…”