µ-Theraphotoxin-Pn3a inhibition of CaV3.3 channels reveals a novel isoform-selective drug binding site

Abstract
Data availability
Article and author information
Metrics

Abstract

Low voltage-activated calcium currents are mediated by T-type calcium channels Ca_V3.1, Ca_V3.2, and Ca_V3.3, which modulate a variety of physiological processes including sleep, cardiac pace-making, pain, and epilepsy. Ca_V3 isoforms' biophysical properties, overlapping expression, and lack of subtype-selective pharmacology hinder the determination of their specific physiological roles in health and disease. We have identified μ-theraphotoxin Pn3a as the first subtype-selective spider venom peptide inhibitor of Ca_V3.3, with >100-fold lower potency against the other T-type isoforms. Pn3a modifies Ca_V3.3 gating through a depolarizing shift in the voltage dependence of activation thus decreasing Ca_V3.3-mediated currents in the normal range of activation potentials. Paddle chimeras of K_V1.7 channels bearing voltage sensor sequences from all four Ca_V3.3 domains revealed preferential binding of Pn3a to the S3-S4 region of domain II (Ca_V3.3^DII). This novel T-type channel pharmacological site was explored through computational docking simulations of Pn3a, site-directed mutagenesis, and full domain II swaps between Cav3 channels highlighting it as a subtype-specific pharmacophore. This research expands our understanding of T-type calcium channel pharmacology and supports the suitability of Pn3a as a molecular tool in the study of the physiological roles of Ca_V3.3 channels.

Data availability

All data generated or analysed during this study are included in the manuscript and supporting file

Article and author information

Author details

Jeffrey R McArthur

Illawarra Health and Medical Research Institute, University of Wollongong, Wollongong, Australia

For correspondence
jeffreym@uow.edu.au

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-2546-7913
Jierong Wen

School of Science, RMIT University, Melbourne, Australia

Competing interests
The authors declare that no competing interests exist.
Andrew Hung

School of Science, RMIT University, Melbourne, Australia

Competing interests
The authors declare that no competing interests exist.
Rocio K Finol-Urdaneta

Illawarra Health and Medical Research Institute, University of Wollongong, Wollongong, Australia

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0003-2157-4532
David J Adams

Illawarra Health and Medical Research Institute, University of Wollongong, Wollongong, Australia

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-7030-2288

Funding

Rebecca L. Cooper Medical Research Foundation (PG2019396)

Jeffrey R McArthur

National Health and Medical Research Council (APP1072113)

David J Adams

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.