An alternative mechanism for the methylation of phosphoethanolamine catalyzed by Plasmodium falciparum phosphoethanolamine methyltransferase

J Biol Chem. 2014 Dec 5;289(49):33815-25. doi: 10.1074/jbc.M114.611319. Epub 2014 Oct 6.


The phosphobase methylation pathway catalyzed by the phosphoethanolamine methyltransferase in Plasmodium falciparum (PfPMT), the malaria parasite, offers an attractive target for anti-parasitic drug development. PfPMT methylates phosphoethanolamine (pEA) to phosphocholine for use in membrane biogenesis. Quantum mechanics and molecular mechanics (QM/MM) calculations tested the proposed reaction mechanism for methylation of pEA involving the previously identified Tyr-19-His-132 dyad, which indicated an energetically unfavorable mechanism. Instead, the QM/MM calculations suggested an alternative mechanism involving Asp-128. The reaction coordinate involves the stepwise transfer of a proton to Asp-128 via a bridging water molecule followed by a typical Sn2-type methyl transfer from S-adenosylmethionine to pEA. Functional analysis of the D128A, D128E, D128Q, and D128N PfPMT mutants shows a loss of activity with pEA but not with the final substrate of the methylation pathway. X-ray crystal structures of the PfPMT-D128A mutant in complex with S-adenosylhomocysteine and either pEA or phosphocholine reveal how mutation of Asp-128 disrupts a hydrogen bond network in the active site. The combined QM/MM, biochemical, and structural studies identify a key role for Asp-128 in the initial step of the phosphobase methylation pathway in Plasmodium and provide molecular insight on the evolution of multiple activities in the active site of the PMT.

Keywords: Computer Modeling; Crystallography; Enzyme Mechanism; Mutagenesis; Protein Structure.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Aspartic Acid / chemistry*
  • Aspartic Acid / metabolism
  • Biocatalysis
  • Crystallography, X-Ray
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Ethanolamines / chemistry*
  • Ethanolamines / metabolism
  • Evolution, Molecular
  • Gene Expression
  • Kinetics
  • Methylation
  • Methyltransferases / chemistry*
  • Methyltransferases / genetics
  • Methyltransferases / metabolism
  • Models, Molecular
  • Phylogeny
  • Plasmodium falciparum / chemistry*
  • Plasmodium falciparum / enzymology
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Protozoan Proteins / chemistry*
  • Protozoan Proteins / genetics
  • Protozoan Proteins / metabolism
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Thermodynamics


  • Ethanolamines
  • Protozoan Proteins
  • Recombinant Proteins
  • Aspartic Acid
  • phosphorylethanolamine
  • Methyltransferases
  • phosphoethanolamine methyltransferase

Associated data

  • PDB/3UJ7
  • PDB/3UJB
  • PDB/4R6W
  • PDB/4R6X