Expression, purification, and characterization of anuran saxiphilins using thermofluor, fluorescence polarization, and isothermal titration calorimetry.

Anuran saxiphilins (Sxphs) are "toxin sponge" proteins thought to prevent the lethal effects of small-molecule neurotoxins through sequestration. Here, we present a protocol for the expression, purification, and characterization of Sxphs. We describe steps for using thermofluor, fluorescence polarization, and isothermal titration calorimetry assays that probe Sxph:saxitoxin interactions using a range of sample quantities. These assays are generalizable and can be used for other paralytic shellfish poisoning toxin-binding proteins. For complete details on the use and execution of this protocol, please refer to Chen et al. (2022). ¹ .
Competing Interests: Declaration of interests J.D.B. is a cofounder and holds equity shares in SiteOne Therapeutics, Inc., a start-up company interested in developing subtype-selective modulators of sodium channels. Patent applications have been filed for this work.
(Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)