Site-Specific Chemoenzymatic Labeling of Aerolysin Enables the Identification of New Aerolysin Receptors
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Wuethrich-2014-Site-specific chemoe.pdf
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Author(s)
Wuethrich, Irene
Peeters, Janneke G. C.
Blom, Annet E. M.
Theile, Christopher S.
Li, Zeyang
Spooner, Eric
Ploegh, Hidde
Guimaraes, Carla P.
Date Issued
October 2014
Journal
PLoS ONE
Publisher
Public Library of Science
Citation
Wuethrich, Irene, Janneke G. C. Peeters, Annet E. M. Blom, Christopher S. Theile, Zeyang Li, Eric Spooner, Hidde L. Ploegh, and Carla P. Guimaraes. “Site-Specific Chemoenzymatic Labeling of Aerolysin Enables the Identification of New Aerolysin Receptors.” Edited by Ludger Johannes. PLoS ONE 9, no. 10 (October 2, 2014): e109883.
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Final published version
Abstract
Aerolysin is a secreted bacterial toxin that perforates the plasma membrane of a target cell with lethal consequences. Previously explored native and epitope-tagged forms of the toxin do not allow site-specific modification of the mature toxin with a probe of choice. We explore sortase-mediated transpeptidation reactions (sortagging) to install fluorophores and biotin at three distinct sites in aerolysin, without impairing binding of the toxin to the cell membrane and with minimal impact on toxicity. Using a version of aerolysin labeled with different fluorophores at two distinct sites we followed the fate of the C-terminal peptide independently from the N-terminal part of the toxin, and show its loss in the course of intoxication. Making use of the biotinylated version of aerolysin, we identify mesothelin, urokinase plasminogen activator surface receptor (uPAR, CD87), glypican-1, and CD59 glycoprotein as aerolysin receptors, all predicted or known to be modified with a glycosylphosphatidylinositol anchor. The sortase-mediated reactions reported here can be readily extended to other pore forming proteins.
MIT Department
Massachusetts Institute of Technology. Department of Biology
Massachusetts Institute of Technology. Department of Biology
Massachusetts Institute of Technology. Department of Chemistry
Whitehead Institute for Biomedical Research
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DOI of Published Version
https://doi.org/10.1371/journal.pone.0109883
