Department of Chemistry and Institutes of Biomedical Sciences, Fudan University, Shanghai 200433, People's Republic of China.
Protein Sci. 2010 Nov;19(11):2073-84. doi: 10.1002/pro.487.
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.
我们成功地从微小隐孢子虫（Cryptosporidium parvum）中表达了重组的线粒体型铁氧还蛋白（mtFd）和铁氧还蛋白：NADP(+)还原酶（mtFNR），并首次对其生物化学特性进行了描述，这是一种顶复门生物。我们获得并纯化了微小隐孢子虫 mtFd（CpmtFd）和 FNR（CpmtFNR）的全蛋白，通过紫外/可见分光光度法和电子顺磁共振证实并表征了 Fd 中[2Fe-2S]簇和 FNR 中 FAD 的正确组装。这些蛋白具有完全的功能，并且在细胞色素 c 偶联测定中，CpmtFNR 能够将电子从 NADPH 转移到 CpmtFd，遵循典型的米氏动力学。顶复门生物的 mtFd 和 mtFNR 蛋白在进化上与人类和动物中的对应蛋白有很大的差异，因此可以作为微小隐孢子虫和其他顶复门生物的潜在药物靶点进行探索。
