Engineering resistance to 'aging' of phosphylated human acetylcholinesterase. Role of hydrogen bond network in the active center

FEBS Lett. 1993 Nov 15;334(2):215-20. doi: 10.1016/0014-5793(93)81714-b.


Recombinant human acetylcholinesterase (HuAChE) and selected mutants (E202Q, Y337A, E450A) were studied with respect to catalytic activity towards charged and noncharged substrates, phosphylation with organophosphorus (OP) inhibitors and subsequent aging of the OP-conjugates. Amino acid E450, unlike residues E202 and Y337, is not within interaction distance from the active center. Yet, the bimolecular rates of catalysis and phosphylation are 30-100 fold lower for both E450A and E202Q compared to Y337A or the wild type and in both mutants the resulting OP-conjugates show striking resistance to aging. It is proposed that a hydrogen bond network, that maintains the functional architecture of the active center, involving water molecules and residues E202 and E450, is responsible for the observed behaviour.

Publication types

  • Comparative Study
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Acetylcholinesterase / chemistry*
  • Acetylcholinesterase / metabolism*
  • Amino Acid Sequence
  • Binding Sites
  • Cholinesterase Inhibitors / pharmacology*
  • Enzyme Stability
  • Humans
  • Hydrogen Bonding
  • Isoflurophate / pharmacology
  • Kinetics
  • Mutagenesis, Site-Directed
  • Phosphorylation
  • Point Mutation
  • Protein Conformation
  • Protein Engineering
  • Quinolinium Compounds / pharmacology
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • Soman / pharmacology


  • Cholinesterase Inhibitors
  • Quinolinium Compounds
  • Recombinant Proteins
  • Isoflurophate
  • 7-((methylethoxyphosphinyl)oxy)-1-methylquinolinium
  • Soman
  • Acetylcholinesterase