title: Cysteine-3 and cysteine-4 are essential for the thioredoxin-like oxidoreductase and antioxidant activities of Plasmodium falciparum macrophage migration inhibitory factor.
abstract: Plasmodium falciparum macrophage migration inhibitory factor (PfMIF) exhibits thioredoxin (Trx)-like oxidoreductase activity but the active site for this activity and its function have not been evaluated. A bioinformatics search revealed that the conserved CXXC motif, which is responsible for Trx-like oxidoreductase activity, is absent from PfMIF. In contrast, the adjacent N-terminal Cys-3 and Cys-4 are conserved in MIF across species of malarial parasites. Mutation of either vicinal Cys-3 or Cys-4 of PfMIF abolished the Trx-like activity, whereas the mutation of the remaining Cys-59 or Cys-103 did not affect it. PfMIF has an antioxidant function. It prevents reactive oxygen species-mediated lipid peroxidation and oxidative damage of DNA as evident from DNA nicking assay. Interestingly, chemical modification of the vicinal cysteines by phenylarsine oxide (PAO), a specific vicinal thiol modifier, significantly prevented this antioxidant activity. Modification of Cys-3 and Cys-4 was confirmed by MALDI-TOF mass spectroscopy of peptide fragments obtained after cyanogen bromide digestion of PAO-modified PfMIF. Furthermore, mutation of either Cys-3 or Cys-4 of PfMIF resulted in the loss of both Trx-like oxidoreductase and antioxidant activities of PfMIF. Altogether, our results suggest that the vicinal Cys-3 and Cys-4 play a critical role in the Trx-like oxidoreductase activity and antioxidant property of PfMIF.
authors.affiliation: Department of Infectious Diseases and Immunology, Indian Institute of Chemical Biology, Jadavpur, Kolkata 700032, West Bengal, India.
authors.forename: Athar
authors.initials: A
authors.lastname: Alam
authors.forename: Manish
authors.initials: M
authors.lastname: Goyal
authors.forename: Mohd Shameel
authors.initials: MS
authors.lastname: Iqbal
authors.forename: Samik
authors.initials: S
authors.lastname: Bindu
authors.forename: Sumanta
authors.initials: S
authors.lastname: Dey
authors.forename: Chinmay
authors.initials: C
authors.lastname: Pal
authors.forename: Pallab
authors.initials: P
authors.lastname: Maity
authors.forename: Nahren Manuel
authors.initials: NM
authors.lastname: Mascarenhas
authors.forename: Nanda
authors.initials: N
authors.lastname: Ghoshal
authors.forename: Uday
authors.initials: U
authors.lastname: Bandyopadhyay
mesh_terms: D000818:Animals; D001152:Arsenicals / Q000494:pharmacology; D019295:Computational Biology; D003545:Cysteine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004249:DNA Damage / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D004912:Erythrocytes / Q000378:metabolism* / Q000469:parasitology / Q000473:pathology; D008263:Macrophage Migration-Inhibitory Factors / Q000737:chemistry / Q000378:metabolism*; D008288:Malaria / Q000235:genetics / Q000378:metabolism* / Q000469:parasitology; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics; D018384:Oxidative Stress / Q000187:drug effects; D010088:Oxidoreductases / Q000378:metabolism; D010963:Plasmodium falciparum / Q000201:enzymology* / Q000472:pathogenicity; D015800:Protozoan Proteins / Q000737:chemistry / Q000378:metabolism*; D011817:Rabbits; D013237:Stereoisomerism; D013879:Thioredoxins / Q000737:chemistry / Q000378:metabolism
chemical_list: D001152:Arsenicals; D008263:Macrophage Migration-Inhibitory Factors; D015800:Protozoan Proteins; C519265:macrophage-migration inhibitory factor, Plasmodium falciparum; C029341:oxophenylarsine; D013879:Thioredoxins; D010088:Oxidoreductases; D003545:Cysteine