Direct tests of enzymatic heme degradation by the malaria parasite Plasmodium falciparum

J Biol Chem. 2012 Nov 2;287(45):37793-807. doi: 10.1074/jbc.M112.414078. Epub 2012 Sep 19.


Malaria parasites generate vast quantities of heme during blood stage infection via hemoglobin digestion and limited de novo biosynthesis, but it remains unclear if parasites metabolize heme for utilization or disposal. Recent in vitro experiments with a heme oxygenase (HO)-like protein from Plasmodium falciparum suggested that parasites may enzymatically degrade some heme to the canonical HO product, biliverdin (BV), or its downstream metabolite, bilirubin (BR). To directly test for BV and BR production by P. falciparum parasites, we DMSO-extracted equal numbers of infected and uninfected erythrocytes and developed a sensitive LC-MS/MS assay to quantify these tetrapyrroles. We found comparable low levels of BV and BR in both samples, suggesting the absence of HO activity in parasites. We further tested live parasites by targeted expression of a fluorescent BV-binding protein within the parasite cytosol, mitochondrion, and plant-like plastid. This probe could detect exogenously added BV but gave no signal indicative of endogenous BV production within parasites. Finally, we recombinantly expressed and tested the proposed heme degrading activity of the HO-like protein, PfHO. Although PfHO bound heme and protoporphyrin IX with modest affinity, it did not catalyze heme degradation in vivo within bacteria or in vitro in UV absorbance and HPLC assays. These observations are consistent with PfHO's lack of a heme-coordinating His residue and suggest an alternative function within parasites. We conclude that P. falciparum parasites lack a canonical HO pathway for heme degradation and thus rely fully on alternative mechanisms for heme detoxification and iron acquisition during blood stage infection.

Publication types

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

MeSH terms

  • Amino Acid Sequence
  • Bilirubin / metabolism
  • Biliverdine / metabolism
  • Chromatography, High Pressure Liquid
  • Erythrocytes / metabolism
  • Erythrocytes / parasitology
  • Heme / metabolism*
  • Heme Oxygenase (Decyclizing) / genetics
  • Heme Oxygenase (Decyclizing) / metabolism*
  • Humans
  • Malaria, Falciparum / blood
  • Malaria, Falciparum / parasitology
  • Microscopy, Fluorescence
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Plasmodium falciparum / enzymology*
  • Plasmodium falciparum / genetics
  • Plasmodium falciparum / metabolism
  • Protein Binding
  • Proteolysis
  • Protoporphyrins / metabolism
  • Protozoan Proteins / genetics
  • Protozoan Proteins / metabolism*
  • Sequence Homology, Amino Acid
  • Spectrometry, Fluorescence
  • Tandem Mass Spectrometry


  • Protoporphyrins
  • Protozoan Proteins
  • Heme
  • protoporphyrin IX
  • Heme Oxygenase (Decyclizing)
  • Biliverdine
  • Bilirubin