Aging of cholinesterases phosphylated by tabun proceeds through O-dealkylation

Eugénie Carletti, He Li, Bin Li, Fredrik Ekström, Yvain Nicolet, Mélanie Loiodice, Emilie Gillon, Marie T. Froment, Oksana Lockridge, Lawrence M Schopfer, Patrick Masson, Florian Nachon

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Abstract

Human butyrylcholinesterase (hBChE) hydrolyzes or scavenges a wide range of toxic esters, including heroin, cocaine, carbamate pesticides, organophosphorus pesticides, and nerve agents. Organophosphates (OPs) exert their acute toxicity through inhibition of acetylcholinesterase (AChE) by phosphylation of the catalytic serine. Phosphylated cholinesterase (ChE) can undergo a spontaneous, time-dependent process called "aging", during which the OP-ChE conjugate is dealkylated. This leads to irreversible inhibition of the enzyme. The inhibition of ChEs by tabun and the subsequent aging reaction are of particular interest, because tabun-ChE conjugates display an extraordinary resistance toward most current oxime reactivators. We investigated the structural basis of oxime resistance for phosphoramidated ChE conjugates by determining the crystal structures of the non-aged and aged forms of hBChE inhibited by tabun, and by updating the refinement of non-aged and aged tabun-inhibited mouse AChE (mAChE). Structures for non-aged and aged tabun-hBChE were refined to 2.3 and 2.1 Å, respectively. The refined structures of aged ChE conjugates clearly show that the aging reaction proceeds through O-dealkylation of the P(R) enantiomer of tabun. After dealkylation, the negatively charged oxygen forms a strong salt bridge with protonated His438Nε2 that prevents reactivation. Mass spectrometric analysis of the aged tabun-inhibited hBChE showed that both the dimethylamine and ethoxy side chains were missing from the phosphorus. Loss of the ethoxy is consistent with the crystallography results. Loss of the dimethylamine is consistent with acid-catalyzed deamidation during the preparation of the aged adduct for mass spectrometry. The reported 3D data will help in the design of new oximes capable of reactivating tabun-ChE conjugates.

Original languageEnglish (US)
Pages (from-to)16011-16020
Number of pages10
JournalJournal of the American Chemical Society
Volume130
Issue number47
DOIs
StatePublished - Nov 26 2008

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ASJC Scopus subject areas

  • Catalysis
  • Chemistry(all)
  • Biochemistry
  • Colloid and Surface Chemistry

Cite this

Carletti, E., Li, H., Li, B., Ekström, F., Nicolet, Y., Loiodice, M., Gillon, E., Froment, M. T., Lockridge, O., Schopfer, L. M., Masson, P., & Nachon, F. (2008). Aging of cholinesterases phosphylated by tabun proceeds through O-dealkylation. Journal of the American Chemical Society, 130(47), 16011-16020. https://doi.org/10.1021/ja804941z