Identification and characterization of heme-interacting proteins in the malaria parasite, Plasmodium falciparum
Identification and characterization of heme-interacting proteins in the malaria parasite, Plasmodium falciparum
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DOI：
10.1074/jbc.m303634200
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发表时间：
2003-07-25
影响因子：
4.8
通讯作者：
Tilley, L
中科院分区：
文献类型：
--
作者：
Campanale, N;Nickel, C;Tilley, L
血红蛋白被疟疾寄生虫恶性疟原虫降解，产生游离的铁原卟啉IX（FP）作为有毒副产物。在FP结合药物如氯喹存在下，FP解毒被抑制，游离FP的积累被认为是杀死寄生虫的关键机制。在努力确定寄生虫蛋白，可能优先与FP相互作用，我们已经使用了质谱法。与固定在琼脂糖上的FP结合的蛋白质包括恶性疟原虫甘油醛-3-磷酸脱氢酶（PfGAPDH）、恶性疟原虫谷胱甘肽还原酶（PfGR）和恶性疟原虫蛋白质二硫键异构酶。为了检查FP结合的潜在后果，我们检查了FP抑制来自恶性疟原虫和其他来源的GAPDH和GR活性的能力。FP抑制PfGAPDH的酶活性，Ki值为0.2 μ M，而红细胞GAPDH的敏感性低得多。相比之下，PfGR比其人类对应物对FP抑制更具抗性（Ki> 25 μ M）。我们还检测了FP抑制另外的抗氧化酶的活性的能力，所述抗氧化酶是恶性疟原虫硫氧还蛋白还原酶，其表现出1 μ M的Ki值，以及恶性疟原虫谷氧还蛋白，其显示出对FP更适度的敏感性。PfGAPDH对FP的敏感性可能表明寄生虫的糖酵解途径对FP应激的调节特别敏感。该途径的抑制可以驱动通过戊糖磷酸途径的通量，确保足够的还原当量的产生以抵消由FP积聚诱导的氧化应激。
The degradation of hemoglobin by the malaria parasite, Plasmodium falciparum, produces free ferriprotoporphyrin IX ( FP) as a toxic by-product. In the presence of FP-binding drugs such as chloroquine, FP detoxification is inhibited, and the build-up of free FP is thought to be a key mechanism in parasite killing. In an effort to identify parasite proteins that might interact preferentially with FP, we have used a mass spectrometry approach. Proteins that bind to FP immobilized on agarose include P. falciparum glyceraldehyde-3-phosphate dehydrogenase (PfGAPDH), P. falciparum glutathione reductase (PfGR), and P. falciparum protein disulfide isomerase. To examine the potential consequences of FP binding, we have examined the ability of FP to inhibit the activities of GAPDH and GR from P. falciparum and other sources. FP inhibits the enzymic activity of PfGAPDH with a K-i value of 0.2 muM, whereas red blood cell GAPDH is much less sensitive. By contrast, PfGR is more resistant to FP inhibition (K-i > 25 muM) than its human counterpart. We also examined the ability of FP to inhibit the activities of the additional antioxidant enzymes, P. falciparum thioredoxin reductase, which exhibits a K-i value of 1 muM, and P. falciparum glutaredoxin, which shows more moderate sensitivity to FP. The exquisite sensitivity of PfGAPDH to FP may indicate that the glycolytic pathway of the parasite is particularly susceptible to modulation by FP stress. Inhibition of this pathway may drive flux through the pentose phosphate pathway ensuring sufficient production of reducing equivalents to counteract the oxidative stress induced by FP build-up.