Role of cysteines in Plasmodium falciparum circumsporozoite protein: interactions with heparin can rejuvenate inactive protein mutants.
Various pathogenic bacteria, viruses, and protozoan bind to glycosaminoglycan-based receptors on host cells and initiate an infection. Sporozoites of Plasmodium predominantly express circumsporozoite (CS) protein on their surface, which binds to heparan sulfate proteoglycans on liver cell surface that subsequently leads to malaria. Here we show that the interaction of free heparin with this parasite ligand has the potential to be a critical component of invasion. CS protein of P. falciparum contains four cysteines at positions 361, 365, 396, and 401. In this study, all four cysteine residues were mutagenized to alanine both individually and in different combinations. Conversion of cysteine 396 to alanine (protein CS3) led to a 10-fold increase in the binding activity of the protein to HepG2 cells. Replacement of cysteines at positions 361, 365, and 401 either alone or in different combinations led to a near total loss of binding. Surprisingly, activity in these inactive mutants could be effectively restored in the presence of submolar concentrations of heparin. Heparin also up-regulated binding of CS3 at submolar concentrations with respect to the protein but down-regulated binding when present in excess. Given the significantly different concentrations of heparin in different organs of the host and the in vitro results described here one can consider in vivo ramifications of this phenomenon for pathogen targeting of specific organs and for the functional effects of antigenic variation on receptor ligand interaction.
Rathore
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Growth and Development Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 9000 Rockville Pike, Bethesda, MD 20892-0425, USA.
McCutchan
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TF
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001667:Binding, Competitive; D003545:Cysteine / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D015870:Gene Expression; D006493:Heparin / Q000378:metabolism*; D019763:Heparin Lyase / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010455:Peptides / Q000378:metabolism; D010963:Plasmodium falciparum; D015800:Protozoan Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism / Q000502:physiology*; D014407:Tumor Cells, Cultured
D010455:Peptides; D015800:Protozoan Proteins; C041595:circumsporozoite protein, Protozoan; D006493:Heparin; D019763:Heparin Lyase; D003545:Cysteine