Substrate-Induced Unfolding of Protein Disulfide Isomerase Displaces the Cholera Toxin A1 Subunit from its Holotoxin

Michael Taylor,Tuhina Banerjee,Supriyo Ray,Helen Burress,Suren A Tatulian,Ken Teter

doi:10.1016/j.bpj.2013.11.2668

Michael Taylor, Tuhina Banerjee + Show 4 more

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https://doi.org/10.1016/j.bpj.2013.11.2668

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Journal: Biophysical journal	Publication Date: Jan 1, 2014
Citations: 50	License type: publisher-specific-oa

Affiliation: University of Central Florida

Abstract

To generate a cytopathic effect, the catalytic A1 subunit of cholera toxin (CT) must be separated from the rest of the toxin. Protein disulfide isomerase (PDI) is thought to mediate CT disassembly by acting as a redox-driven chaperone that actively unfolds the CTA1 subunit. Here, we show by isotope-edited Fourier transform infrared spectroscopy and circular dichroism that PDI itself unfolds upon contact with CTA1. The substrate-induced unfolding of PDI provides a novel molecular mechanism for holotoxin disassembly: we postulate the expanded hydrodynamic radius of unfolded PDI acts as a wedge to dislodge reduced CTA1 from its holotoxin. The oxidoreductase activity of PDI was not required for CT disassembly, but CTA1 displacement did not occur when PDI was locked in a folded conformation or when its substrate-induced unfolding was blocked due to the loss of chaperone function. Our data establish a new property of PDI that is required for cholera intoxication and may be linked to its function as a chaperone that prevents protein aggregation.

Highlights

2386-Pos Board B78 Regulation of Disulfide Coupled Folding of De Novo Designed Precursor Protein Shiori Fukumoto1, Yuichiro Yoshida1, Takuma Maekawa2, Masaki Okumura3, Hiroshi Yamaguchi2, Shigeru Shimamoto1, Yuji Hidaka1. 1Kinki University, Higashi-osaka, Japan, 2Kwansei Gakuin University, Sanda, Japan, 3Tohoku University, Sendai, Japan
Mis-bridged disulfide isomers that possess relatively long half-lives expose their hydrophobic surfaces to solvents, resulting in non-specific aggregation during folding. To overcome this issue and investigate the mechanism of disulfide-coupled protein folding, a series of thiol reagents was prepared and their ability to regulate the folding of disulfide-containing proteins was examined.We recently reported that Arg-Cys-Gly (RCG) accelerated the folding of disulfide-containing-proteins and proposed that positivelycharged redox reagents are effective in promoting disulfide-coupled protein folding
Uroguanylin, a peptide consisting of 16 amino acid residues and 2 disulfide bonds, participates in salt and water homeostasis in mammals and the correct disulfide pairings are absolutely required for its biological activity

Summary

Introduction

2386-Pos Board B78 Regulation of Disulfide Coupled Folding of De Novo Designed Precursor Protein Shiori Fukumoto1, Yuichiro Yoshida1, Takuma Maekawa2, Masaki Okumura3, Hiroshi Yamaguchi2, Shigeru Shimamoto1, Yuji Hidaka1. 1Kinki University, Higashi-osaka, Japan, 2Kwansei Gakuin University, Sanda, Japan, 3Tohoku University, Sendai, Japan. Protein disulfide isomerase (PDI) is thought to mediate CT disassembly by acting as a redox-driven chaperone that actively unfolds the CTA1 subunit. The oxidoreductase activity of PDI was not required for CT disassembly, but CTA1 displacement did not occur when PDI was locked in a folded conformation or when its substrate-induced unfolding was blocked due to the loss of chaperone function.

Results

Conclusion