α-Conotoxin Decontamination Protocol Evaluation: What Works and What Doesn't.

Matthew Turner,Owen Mcdougal,John Cort

doi:10.3390/toxins9090281

Matthew Turner, Owen Mcdougal + Show 1 more

Open Access

https://doi.org/10.3390/toxins9090281

Copy DOI

Abstract

Nine publically available biosafety protocols for safely handling conotoxin peptides were tested to evaluate their decontamination efficacy. Circular dichroism (CD) spectroscopy and mass spectrometry (MS) were used to assess the effect of each chemical treatment on the secondary and primary structure of α-CTx MII (L10V, E11A). Of the nine decontamination methods tested, treatment with 1% (m/v) solution of the enzymatic detergent Contrex™ EZ resulted in a 76.8% decrease in α-helical content as assessed by the mean residue ellipticity at 222 nm, and partial peptide digestion was demonstrated using high performance liquid chromatography mass spectrometry (HPLC-MS). Additionally, treatment with 6% sodium hypochlorite (m/v) resulted in 80.5% decrease in α-helical content and complete digestion of the peptide. The Contrex™ EZ treatment was repeated with three additional α-conotoxins (α-CTxs), α-CTxs LvIA, ImI and PeIA, which verified the decontamination method was reasonably robust. These results support the use of either 1% Contrex™ EZ solution or 6% sodium hypochlorite in biosafety protocols for the decontamination of α-CTxs in research laboratories.

Highlights

The venom of predatory marine snails from the genus Conus is comprised of as many as 1000 distinct neurologically active peptides [1]
To determine the effect of chemical treatment on the secondary structure of α-CTx MII (L10V, E11A), α-helical content was estimated from the measured ellipticity at 222 nm
Near-UV Circular dichroism (CD) spectroscopy indicated that DTT effectively reduced the disulfide bonds of α-CTx

Summary

Introduction

The venom of predatory marine snails from the genus Conus is comprised of as many as 1000 distinct neurologically active peptides [1]. These small disulfide rich peptide toxins are referred to as conotoxins (CTxs). CTxs vary in their mechanism of action, and many have been found to modulate ion channels. These include α-CTxs which inhibit ligand gated nicotinic acetylcholine receptors (nAChRs), δ-CTxs which inhibit inactivation of voltage-dependent sodium channels, ω-CTxs which inhibit N-type voltage-dependent calcium channels, and κ-CTxs which inhibit potassium channels [2]. Because CTxs represent extremely specific molecular probes, they are routinely used by researchers as a tool to study and differentiate between closely related biological receptor subtypes

Objectives

Methods

Results

Conclusion