BPS2026 – Atomic signatures of adaptation: Tetrodotoxin-resistant sodium channels in snakes

Tetrodotoxin (TTX) is a neurotoxin that binds to the pore of voltage-gated sodium (Na V ) channels and inhibits neurocellular communication. Californian populations of garter snakes ( Thamnophis sirtalis ) that prey on TTX-producing newts have Na V channels that are extremely resistant to TTX, a phenotype conferred by four amino acid substitutions relative to sensitive channels. While electrophysiological assays have quantified TTX resistance in these snake populations, an atomistic understanding of this phenotype has not been developed. Here, we investigated the biophysical mechanism for TTX resistance by comparing the dynamics of snake Na V channels to that of a TTX-sensitive cockroach channel. Four systems were simulated using the CHARMM36 force field, each with and without TTX bound: (1) a sensitive cockroach channel, (2) a sensitive snake channel, (3) a resistant snake channel, and (4) a cockroach channel with four resistance-conferring snake mutations. We employed covariance analysis to identify differences in motion due to mutation or TTX binding that may rationalize TTX resistance. We found that the baseline dynamics of the WT cockroach are perturbed by the four resistance-conferring mutations, with most of the mutated region behaving differently and TTX dissociation from the pore in one of the mutated cockroach replicates. When comparing sensitive and resistant snake channels in the presence of TTX, the voltage-sensing domain connected to the pore mutations exhibits different dynamics, which could explain the trade-off in NaV channel activation observed in extremely resistant snakes. The mutations behave the same in the snake and cockroach channels when TTX is present, which could mean those residues may influence overall pore dynamics in the presence of TTX. Further characterization of the difference in dynamics will help determine how natural selection not only acts at the molecular level but fine-tunes the atomistic dynamics within a protein to confer an adaptive molecular phenotype.