Preventing voltage-dependent gating of anthrax toxin channels using engineered disulfides.

The Journal of General Physiology
Damon S Anderson, R O Blaustein

Abstract

The channel-forming component of anthrax toxin, (PA(63))(7), is a heptameric water-soluble protein at neutral pH, but under acidic conditions it spontaneously inserts into lipid bilayers to form a 14-stranded beta-barrel ion-conducting channel. This channel plays a vital role in anthrax pathogenesis because it serves as a conduit for the membrane translocation of the two enzymatic components of anthrax toxin, lethal factor and edema factor. Anthrax channels open and close in response to changes in transmembrane voltage, a property shared by several other pore-forming toxins. We have discovered an unexpected phenomenon in cysteine-substituted channels that provides a window into this gating process: their normal voltage-dependent gating can be abolished by reaction with methanethiosulfonate (MTS) reagents or exposure to oxidizing conditions. Remarkably, this perturbation is seen with cysteines substituted at sites all along the approximately 100 A length of the channel's beta-barrel. In contrast, reaction with N-ethylmaleimide, a thiol-reactive compound that does not form a mixed disulfide, does not affect gating at any of the sites tested. These findings, coupled with our biochemical detection of dimers, have led us to conclude...Continue Reading

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Citations

Oct 13, 2012·Chemical Reviews·Ekaterina M Nestorovich, Sergey M Bezrukov
Mar 16, 2011·The Journal of General Physiology·Robert O Blaustein
Jul 15, 2015·The Journal of General Physiology·Aviva SchiffmillerAlan Finkelstein
Jul 27, 2010·Biophysical Journal·Ekaterina M NestorovichSergey M Bezrukov
Nov 18, 2017·Journal of Molecular Microbiology and Biotechnology·Gabriel Moreno-HagelsiebMilton H Saier
Aug 2, 2021·Biochimica Et Biophysica Acta. Biomembranes·Wenxing Liu, Ekaterina M Nestorovich
Apr 13, 2013·Biochemistry·Indra SchroederAnna Moroni

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Methods Mentioned

BETA
x-ray crystallography

Software Mentioned

PyMol
MTSET
ChemDraw
Axon pClamp

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