Title

Stabilization Of The Tertiary Structure Of The Cholera Toxin A1 Subunit Inhibits Toxin Dislocation And Cellular Intoxication

Keywords

cholera toxin; circular dichroism; endoplasmic reticulum-associated degradation; surface plasmon resonance; toxin translocation

Abstract

Cholera toxin (CT) moves from the cell surface to the endoplasmic reticulum (ER) by retrograde vesicular transport. The catalytic subunit of CT (CTA1) then crosses the ER membrane and enters the cytosol in a process that involves the quality control mechanism of ER-associated degradation. The molecular details of this dislocation event have not been fully characterized. Here, we report that thermal instability in the CTA1 subunit-specifically, the loss of CTA1 tertiary structure at 37 °C-triggers toxin dislocation. Biophysical studies found that glycerol preferentially stabilized the tertiary structure of CTA1 without having any noticeable effect on the thermal stability of its secondary structure. The thermal disordering of CTA1 tertiary structure normally preceded the perturbation of its secondary structure, but in the presence of 10% glycerol the temperature-induced loss of CTA1 tertiary structure occurred at higher temperatures in tandem with the loss of CTA1 secondary structure. The glycerol-induced stabilization of CTA1 tertiary structure blocked CTA1 dislocation from the ER and instead promoted CTA1 secretion into the extracellular medium. This, in turn, inhibited CT intoxication. Glycerol treatment also inhibited the in vitro degradation of CTA1 by the core 20S proteasome. Collectively, these findings indicate that toxin thermal instability plays a key role in the intoxication process. They also suggest the stabilization of CTA1 tertiary structure is a potential goal for novel antitoxin therapeutic agents. © 2009 Elsevier Ltd. All rights reserved.

Publication Date

11-13-2009

Publication Title

Journal of Molecular Biology

Volume

393

Issue

5

Number of Pages

1083-1096

Document Type

Article

Personal Identifier

scopus

DOI Link

https://doi.org/10.1016/j.jmb.2009.09.013

Socpus ID

70350025562 (Scopus)

Source API URL

https://api.elsevier.com/content/abstract/scopus_id/70350025562

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