An Appetite for Destruction: Detecting Prey-Selective Binding of α-Neurotoxins in the Venom of Afro-Asian Elapids.

Richard J Harris,Bryan G Fry,Christina N Zdenek,Nathaniel Frank,David Harrich

doi:10.3390/toxins12030205

Richard J Harris, Bryan G Fry + Show 3 more

Open Access

https://doi.org/10.3390/toxins12030205

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Abstract

Prey-selective venoms and toxins have been documented across only a few species of snakes. The lack of research in this area has been due to the absence of suitably flexible testing platforms. In order to test more species for prey specificity of their venom, we used an innovative taxonomically flexible, high-throughput biolayer interferometry approach to ascertain the relative binding of 29 α-neurotoxic venoms from African and Asian elapid representatives (26 Naja spp., Aspidelaps scutatus, Elapsoidea boulengeri, and four locales of Ophiophagus hannah) to the alpha-1 nicotinic acetylcholine receptor orthosteric (active) site for amphibian, lizard, snake, bird, and rodent targets. Our results detected prey-selective, intraspecific, and geographical differences of α-neurotoxic binding. The results also suggest that crude venom that shows prey selectivity is likely driven by the proportions of prey-specific α-neurotoxins with differential selectivity within the crude venom. Our results also suggest that since the α-neurotoxic prey targeting does not always account for the full dietary breadth of a species, other toxin classes with a different pathophysiological function likely play an equally important role in prey immobilisation of the crude venom depending on the prey type envenomated. The use of this innovative and taxonomically flexible diverse assay in functional venom testing can be key in attempting to understanding the evolution and ecology of α-neurotoxic snake venoms, as well as opening up biochemical and pharmacological avenues to explore other venom effects.

Highlights

The role of predator–prey relationship dynamics plays a key role in our understanding of how prey provide positive selection pressures for toxins and crude venom to become more prey specific.Regarding toxin diversification and evolution, the Red Queen hypothesis [1] would suggest that an arms race between predator and prey would provide a positive selection pressure for predatory venom toxins to evolve prey specificity and, reciprocally, for toxin resistance to evolve in prey and Toxins 2020, 12, 205; doi:10.3390/toxins12030205 www.mdpi.com/journal/toxinsToxins 2020, 12, 205 predators of venomous animals
Snake venoms are predominantly used for predatory purposes; it is likely that these co-evolutionary arms races and selective pressures are acting upon some snake venom toxins to evolve target specificity to immobilise certain prey types more efficiently
Our data indicate that the neurotoxic nicotinic acetylcholine receptors (nAChRs) binding across the majority of Naja tested showed some selectivity toward the amphibian mimotope (Figure 1) with 70% of Naja species binding more selectively to the amphibian, and this target being a close second highest in all five other Naja venoms that did not have amphibian as their highest binding (Figure 1)

Summary

Introduction

The role of predator–prey relationship dynamics plays a key role in our understanding of how prey provide positive selection pressures for toxins and crude venom to become more prey specific. Molecular evidence likewise suggests that some venom toxins are under high positive selection pressure, causing rapid diversification through mutations that alter protein structure and function [14,15,16] These selection pressures have been reportedly due to preferential prey targets driving the selection pressures of venom potency [15,16,17,18]. In order to help our understanding of prey-specific binding of toxins and how these shape overall venom evolution, this study utilised a validated biolayer interferometry (BLI) assay [32] to test a large diversity of 3FTx-rich venoms from 29 African and Asian elapids (26 Naja spp., Aspidelaps scutatus, Elapsoidea boulengeri, and four locales of Ophiophagus hannah) and a 3FTx representative (alpha-cobratoxin) against the alpha-1 nAChR orthosteric sites from some of the major groups of potential prey types: amphibian, lizard, snake, bird, and rodent. It is likely that the overall proportions of prey-specific toxins within the venom play a greater role in crude-venom specificity and that diet range may moderate the overall proportions of prey-specific toxins

Results and Discussion

Venom Collection and Preparation

Mimotope Production and Preparation

Data Processing and Statistical Analyses