Molecular organization of mitochondrial quinol-fumarate reductase and its role in oxygen adaptation.

猪蛔虫成虫肌肉复合体II具有较高的喹啉-延胡索酸还原酶（QFR）活性，在其微氧环境适应过程中的无氧电子传递中起着关键作用。相反，幼虫（L3）复合物II显示出比成人酶低得多的QFR活性，并在有氧呼吸中起琥珀酸泛醌还原酶（SQR）的作用。我们报道了猪蛔虫线粒体复合物II的阶段特异性异构体，并表明至少在幼虫期复合物II的黄素蛋白亚基（Fp）和细胞色素B小亚基（cybS）与成虫期不同。在本研究中，完整的cDNA的铁-硫亚基（Ip）的复合物II，它与Fp形成复合物II的催化部分，已被克隆和测序的厌氧成年猪蛔虫，和自由生活的线虫，秀丽隐杆线虫。这两种线虫的Ip亚基的氨基酸序列是相似的，特别是在三个富含半胱氨酸的区域，这三个区域被认为是共同的。 ...更多信息 撬起酶的铁硫簇。由于北方杂交显示成体Ip也在L3中表达，因此也表征了来自猪蛔虫幼虫的Ip。幼虫复合物II的IP被识别的抗体对成人IP，并从成人IP的肽图谱难以区分。经DEAE-cellulofine柱层析纯化的幼虫复合物II中IP的N-苏氨酸42氨基酸序列与成虫IP的成熟形式相同。此外，在PVDF膜上的微量分析测定的幼虫IP的氨基酸组成几乎是相同的成年IP。这些结果，连同使用简并引物通过RT-PCR的同源性探测未能找到幼虫特异性Ip的事实，表明两种不同阶段特异性形式的猪曲霉复合物II共享一个共同的Ip亚基，即使成人酶作为QFR的功能，而幼虫酶作为SQR.In此外，我们发现了新的化合物，萘呋啶，从尼日尔曲霉。萘呋啶抑制NADH-延胡索酸还原酶（复合物I+II）的活性，一个独特的无氧电子传递系统在hilminth线粒体，在nM的顺序。它竞争复合物I中的醌结合位点，并对蠕虫酶显示出高选择性毒性。此外，萘呋啶在使用绵羊的体内试验中对捻转血矛线虫发挥驱虫活性。因此，我们的研究表明，线粒体复合物I是一个很有前途的化疗靶点，萘呋啶是一个新的潜在的铅化合物作为一种新的驱虫剂从微生物中分离。少

Complex II of adult Ascaris suum muscle exhibits high quinol-fumarate reductase(QFR)activity and plays a key role in anaerobic electron-transport during adaptation to their microaerobic habitat. In contrast, larval(L3)complex II shows a much lower QFR activity than the adult enzyme, and functions as succinate-ubiquinone reductase(SQR)in aerobic respiration. We have reported the stage-specific isoforms of complex II in A.suum mitochondria, and showed that at least the flavoprotein subunit(Fp)and the small subunit of cytochrome b(cybS)of the larval complex II differ from those of adult. In the present study, complete cDNAs for the iron-sulfur subunit(Ip)of complex II, which with Fp forms the catalytic portion of complex II, have been cloned and sequenced from anaerobic adult A.suum, and the free-living nematode, Caenorhabditis elegans. The amino acid sequences of the Ip subunits of these two nematodes are similar, particularly around the three cystein-rich regions that are thought to com … More prise the iron-sulfur clusters of the enzyme. The Ip from A.suum Larvae also was characterized because Northern hybridization showed that the adult Ip also is expressed in L3. The Ip of larval complex II was recognized by the antibody against adult Ip, and was indistinguishable from the adult Ip by peptide mapping. The N-therminal 42 amino acid sequence of Ip in the larval complex II purified by DEAE-cellulofine column chromatography was identical to that of the mature form of the adult Ip. Furthermore, the amino acid composition of larval Ip determined by micro-analysis on a PVDF membrane is almost the same as that of adult Ip. These results, together with the fact that homology probing by RT-PCR using degenerated primers failed to find a larval-specific Ip, suggest that the two different stage-specific forms of the A.suum complex II share a common Ip subunit, even though the adult enzyme functions as a QFR, while larval enzyme acts as an SQR.In addition, we found novel compound, nafuredin, from Aspergillus niger. Nafuredin inhibits NADH-fumarate reductase(complexes I+II)activity, a unique anaerobic electron transport system in hilminth mitochondria, at nM order. It competes for the quinone-binding site in complex I and shows high selective toxicity to the helminth enzyme. Moreover, nafuredin exerts anthelmintic activity against Haemonchus contortus in in vivo trials using sheep. Thus, our study indicates that mitochondrial complex I is a promising target for chemotherapy and that nafuredin is a new potential lead compound as a novel anthelmintic isolated from microorganisms. Less