Title

Contribution Of Subdomain Structure To The Thermal Stability Of The Cholera Toxin A1 Subunit

Abstract

The catalytic A1 subunit of cholera toxin (CTA1) is an ADP- ribosyltransferase with three distinct subdomains: CTA11 forms the catalytic core of the toxin, CTA12 is an extended linker between CTA11 and CTA13, and CTA13 is a compact globular region. CTA1 crosses the endoplasmic reticulum (ER) membrane to enter the cytosol where it initiates a cytopathic effect. Toxin translocation involves ER-associated degradation (ERAD), a quality control system that exports misfolded proteins from the ER to the cytosol. At the physiological temperature of 37 °C, the free CTA1 subunit is in a partially unfolded conformation that triggers its ERAD-mediated translocation to the cytosol. Thus, the temperature sensitivity of CTA1 structure is an important determinant of its function. Here, we examined the contribution of CTA1 subdomain structure to the thermal unfolding of CTA1. Biophysical measurements demonstrated that the CTA1 1 subdomain is thermally unstable and that the CTA12 subdomain provides a degree of conformational stability to CTA11. The CTA13 subdomain does not affect the overall stability of CTA1, but the thermal unfolding of CTA1 appears to begin with a local loss of structure in the CTA13 subdomain: glycerol and acidic pH both inhibited the thermal disordering of full-length CTA1 but not the disordering of a CTA1 construct lacking the A13 subdomain. These observations provide mechanistic insight regarding the thermal unfolding of CTA1, an event which facilitates its subsequent translocation to the cytosol. © 2010 American Chemical Society.

Publication Date

10-19-2010

Publication Title

Biochemistry

Volume

49

Issue

41

Number of Pages

8839-8846

Document Type

Article

Personal Identifier

scopus

DOI Link

https://doi.org/10.1021/bi101201c

Socpus ID

77957907884 (Scopus)

Source API URL

https://api.elsevier.com/content/abstract/scopus_id/77957907884

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