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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.

After forrnation of a ternary complex, SR398/GroESIUbg, cross-linked ubiquitin was discharged by the addition ofa reductant, ]n this systcm, wc have suceeeded in improving the signal-to-noise ratio of the spectrum.

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