Emerging Concepts in Ion Channel Biophysics

Emerging Concepts in Ion Channel Biophysics

Poster Abstracts

58-POS Board 58 New Molecular Determinants of the Voltage-gated Na + Channel Α/Β1 Functional Interface Bertin Paiz-Candia 1 , Thomas Scior 1 , Angel Islas 2,1 , Alfredo Sánchez-Solano 2 , Claudia Mancilla-Simbro 2 , Lourdes Millan-PerezPeña 3 , Eduardo M. Salinas-Stefanon 2 . 1 Facultad de Ciencias Químicas, Universidad Autónoma de Puebla, Puebla, Puebla, Mexico, 2 Laboratorio de Biofísica Instituto de Fisiología, Universidad Autónoma de Puebla, Puebla, Mexico, 3 3Centro de Química, Instituto de Ciencias, Universidad Autónoma de Puebla, Puebla, México, Puebla, Mexico. Most of the inward Na + currents responsible for the initiation of the action potential in vertebrate excitable cells are modulated by membrane immunoglobulin-like (Ig-like) proteins called β1 auxiliary Na+ channel subunits. By the beginning of this century, chimeric studies on the β1- induced electrophysiological modulation of Na + channels had established that it is mediated by an extracellular protein-protein interaction between the pore-forming α subunit and the Ig-like domain of β1. Subsequently, our group has been amidst those who investigate such phenomena at an atomistic level by computational means, i.e. via standard homology molecular modelling and by a novel “analogy approach”. In the absence of the crystal structure of the β1 subunit, we predicted two intramolecular bonds involved in the modulation of the rNav1.4 channel, a disulphide and salt bridge, that were soon later observed in the crystal structure of β3 (a homologue of β1, with a 44% sequence identity). We have also demonstrated that β1 significantly accelerates the inactivation of rNav1.4 channels, in which the intracellular inactivation particle is absent, albeit within seconds. Exhaustive, unbiased sampling and extremely low homology, “analogy modelling”, allowed the identification of two consecutive, key residues on β1, whereby the double mutant: T109A N110A, we called TANA, caused a pervasive electrophysiological loss-of-function. Mutant β1-TANA co-expression with rNav1.4: i) abolished the β1-induced left-hand shift of the steady-state inactivation curve, ii) significantly reduced the β1-induced acceleration of the inactivation and recovery from inactivation, and iii) induced a current rundown upon repetitive depolarizations at 1, 2 and 5 Hz. These results may delineate the functional mammalian Na + channel α/β1 interface and may pave the way for structure-based drug design targeting the β1 subunit, which incidentally also modulates the response to some Na+ channel blockers.

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