Isolation and characterization of the stage-specific cytochrome b small subunit (CybS) of Ascaris suum complex II from the aerobic respiratory chain of larval mitochondria
Isolation and characterization of the stage-specific cytochrome b small subunit (CybS) of Ascaris suum complex II from the aerobic respiratory chain of larval mitochondria
复制标题
DOI：
10.1016/s0166-6851(03)00074-4
复制
发表时间：
2003-05-01
影响因子：
1.5
通讯作者：
Kita, K
中科院分区：
文献类型：
--
作者：
Amino, H;Osanai, A;Kita, K
我们最近报道，猪蛔虫线粒体表达的阶段特异性异构体的复合物11：黄素蛋白亚基和小亚基的细胞色素B（CybS）的幼虫复合物11不同于成人的酶，而两个复合物II共享一个共同的铁硫簇亚基（Ip）。在本研究中，A.猪幼虫复合物II被高度纯化以更详细地表征幼虫细胞色素B亚基。肽质量指纹图谱和N-末端氨基酸序列分析表明，幼虫和成虫的细胞色素B（CybL）蛋白是相同的。相反，cDNA序列显示幼虫细胞色素B的小亚基（CybS（L））不同于成虫细胞色素B的小亚基（CybS（A））。此外，北方分析和免疫印迹显示，阶段特异性表达的CybSL和CybSA在幼虫和成虫线粒体，分别。酶的测定结果表明，rhodoquinol-fumarate还原酶（RQFR）琥珀酸泛醌还原酶（SQR）的活性和醌类的Km值几乎是相同的成人和幼虫复杂IIs的比例，但富马酸还原酶（FRD）的活性是较高的成人形式比幼虫形式。这些结果表明，A.猪复合物II具有与哺乳动物宿主的复合物11不同的性质，并且幼虫复合物11能够起到RQFR的作用。这种幼虫复合物II的RQFR活性对于快速适应宿主感染期间氧可用性的急剧变化是必不可少的。(C)2003 Elsevier Science B. V.保留所有权利。
We recently reported that Ascaris suum mitochondria express stage-specific isoforms of complex 11: the flavoprotein subunit and the small subunit of cytochrome b (CybS) of the larval complex 11 differ from those of adult enzyme, while two complex IIs share a common iron-sulfur cluster subunit (Ip). In the present study, A. suum larval complex II was highly purified to characterize the larval cytochrome b subunits in more detail. Peptide mass fingerprinting and N-terminal amino acid sequencing showed that the larval and adult cytochrome b (CybL) proteins are identical. In contrast, cDNA sequences revealed that the small subunit of larval cytochrome b (CybS(L)) is distinct from the adult CybS (CybS(A)). Furthermore, Northern analysis and immunoblotting showed stage-specific expression of CybSL and CybSA in larval and adult mitochondria, respectively. Enzymatic assays revealed that the ratio of rhodoquinol-fumarate reductase (RQFR) to succinate-ubiquinone reductase (SQR) activities and the K-m values for quinones are almost identical for the adult and larval complex IIs, but that the fumarate reductase (FRD) activity is higher for the adult form than for the larval form. These results indicate that the adult and larval A. suum complex IIs have different properties than the complex 11 of the mammalian host and that the larval complex 11 is able to function as a RQFR. Such RQFR activity of the larval complex II would be essential for rapid adaptation to the dramatic change of oxygen availability during infection of the host. (C) 2003 Elsevier Science B.V. All rights reserved.