The apicomplexan Cryptosporidium parvum possesses a single mitochondrial-type ferredoxin and ferredoxin:NADP+ reductase system.
We have successfully expressed recombinant mitochondrial-type ferredoxin (mtFd) and ferredoxin:NADP(+) reductase (mtFNR) from Cryptosporidium parvum and characterized their biochemical features for the first time for an apicomplexan. Both C. parvum mtFd (CpmtFd) and FNR (CpmtFNR) were obtained and purified as holo-proteins, in which the correct assembly of [2Fe-2S] cluster in Fd and that of FAD in FNR were confirmed and characterized by UV/vis and electron paramagnetic resonance. These proteins were fully functional and CpmtFNR was capable of transferring electrons from NADPH to CpmtFd in a cytochrome c-coupled assay that followed a typical Michaelis-Menten kinetics. Apicomplexan mtFd and mtFNR proteins were evolutionarily divergent from their counterparts in humans and animals and could be explored as potential drug targets in Cryptosporidium and other apicomplexans.
Lei
Cheng
C
Department of Chemistry and Institutes of Biomedical Sciences, Fudan University, Shanghai 200433, People's Republic of China.
Rider
S Dean
SD
Wang
Cai
C
Zhang
Haili
H
Tan
Xiangshi
X
Zhu
Guan
G
D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D016785:Cryptosporidium parvum / Q000201:enzymology* / Q000235:genetics; D004579:Electron Transport; D004591:Electrophoresis, Polyacrylamide Gel; D005287:Ferredoxin-NADP Reductase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D005288:Ferredoxins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020869:Gene Expression Profiling; D006801:Humans; D008018:Life Cycle Stages; D024101:Mitochondrial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D058977:Molecular Sequence Annotation; D008969:Molecular Sequence Data; D010802:Phylogeny; D015800:Protozoan Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D013056:Spectrophotometry, Ultraviolet
D005288:Ferredoxins; D024101:Mitochondrial Proteins; D015800:Protozoan Proteins; D005287:Ferredoxin-NADP Reductase