Electrostatic resistance to alpha-neurotoxins conferred by charge reversal mutations in nicotinic acetylcholine receptors

Abstract

The evolution of venom resistance through coevolutionary chemical arms races has arisen multiple times throughout animalia. Prior documentation of resistance to snake venom α-neurotoxins consists of the N-glycosylation motif or the hypothesized introduction of arginine at positions 187 at the α-1 nicotinic acetylcholine receptor orthosteric site. However, no further studies have investigated the possibility of other potential forms of resistance. Using a biolayer interferometry assay, we first confirm that the previously hypothesized resistance conferred by arginine at position 187 in the honey badger does reduce binding to α-neurotoxins, which has never been functionally tested. We further discovered a novel form of α-neurotoxin resistance conferred by charge reversal mutations, whereby a negatively charged amino acid is replaced by the positively charged amino acid lysine. As venom α-neurotoxins have evolved strong positive charges on their surface to facilitate binding to the negatively charged α-1 orthosteric site, these mutations result in a positive charge/positive charge interaction electrostatically repelling the α-neurotoxins. Such a novel mechanism for resistance has gone completely undiscovered, yet this form of resistance has convergently evolved at least 10 times within snakes. These coevolutionary innovations seem to have arisen through convergent phenotypes to ultimately evolve a similar biophysical mechanism of resistance across snakes.

Widespread convergent evolution of alpha-neurotoxin resistance in African mammals

“…In two of these instances, the Eurasian otter ( Lutra lutra ) and the oryx antelope ( Oryx dammah ), R 187 is not secondarily accompanied by the substitution I 189 or L 189 (Figure 1). Empirical work has shown that nAChR loses affinity for αNTX with the R 187 mutation alone, and that addition of the I 189 , or I 189 by itself does not confer resistance despite its propensity to be paired with R 187 [19]. However, the presence of the single mutant R 187 in these data suggests that the accompanying mutation I 189 is likely not a necessary epistatic mutation and raises the possibility that it may have some function in resistance that is apparent in vivo that is not recovered in vitro [19].…”

Monkeying around with venom: an increased resistance to α-neurotoxins supports an evolutionary arms race between Afro-Asian primates and sympatric cobras

“…Thus, given that commonly encountered neurotoxic snakes exert a strong selection pressure in their defensive envenomations, why does there only seem to be an increased resistance rather than full resistance? Firstly, our aforementioned proposition that there might be an evolutionary trade-off disadvantage to evolving full resistance at the nAChR orthosteric site [ 53 , 60 ] might mean that evolving partial resistance could allow for a balance between reduced α-neurotoxin susceptibility and a somewhat efficient ACh binding. This is supported by previous work revealing that increased resistance toward the nAChR agonist epibatidine decreased ACh sensitivity to the site [ 53 ].…”

Convergent evolution of toxin resistance in animals

“…2; Khan et al ., 2020). Resistance to α‐neurotoxins is less common in non‐elapid neurotoxic snakes – but is ubiquitous within the elapids (Khan et al ., 2020; Harris & Fry, 2021), perhaps suggesting a strong selection pressure for the evolution of autoresistance.…”