Structural and functional analysis of Ascaris suum cytochrome b in the nematode methemoglobin reductase system

线虫高铁血红蛋白还原酶系统中猪蛔虫细胞色素b的结构和功能分析

基本信息

批准号：
18590406
负责人：
TAKAMIYA Shinzaburo
金额：
$ 2.47万
依托单位：
Juntendo University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18590406/
关键词：
cytochrome b_5 methemoglobin reductase system Ascaris suum X-ray crystallography adaptation to low-oxygen tension C. elegans 蛋白質 X-線構造解析結晶化ヘモグロビン

项目摘要

Objectives : This research project was undertaken to elucidate structural properties of Ascaris suum cytochrome b_5, the component of novel NADH-methemoglobin reductase system , by comparing the coding gene and crystal structure with those of cytochromes b_5 from mammalian host and free-living nematode C. elegans. More specifically, 1) To resolve structural properties of A. suum cytochrome b_5 , which have been gained in the course of parasitic adaptation, 2) To analyze its mode of biosynthesis, i. e. the role of presequence of A. suum cytochrome b_5 by expressing the precursor cytochrome b_5 in C. elegans nematode. Achievements 1) The crystal structure of A. suum cytochrome b_5 was resolved at 1.8 A resolution demonstrating the structure different from that of aerobic mammalian cytochrome b_5(erythrocyte type, soluble). Docking models of A. suum hemoglobin and cytochrome b_5 strongly suggest that they are physiological reaction partners. Immunohistochemical and immunoblotting studies … More showed that A. suum cytochrome b_5 was localized in both the hypodermis and perienteric fluid , and that the cytochrome was a secretary protein. We proposed a working hypothesis that the cytochrome b_5 was a protein specialized during adaptation to low-oxygen tension of host intestinal lumen. To test this hypothesis, cytochrome b_5 species were surveyed from gene data bases of C. elegans, of which genome project has been completed. Four species of C. elegans cytochrome _b5 were found although none of them possesses presequence at all. Hydropathy analysis of the four species suggested that two of them were soluble proteins and one of the two exhibited highest homology with A. suum cytochrome b_5. However, none of the two was detected as expressed sequence tag. 2) Experimental conditions are currently examined because of low efficiency of transfection. 3) Affinity analysis using Biacore, employing Ascaris suum and human cytochromes b_5 as ligand, showed that the latter had less affinity with perienteric hemoglobin than the former. These results supported the working hypothesis described above. Less

目的：本研究项目旨在通过将编码基因和晶体结构与来自哺乳动物宿主和自由生活线虫 C 的细胞色素 b_5 的编码基因和晶体结构进行比较，阐明猪蛔虫细胞色素 b_5（新型 NADH-高铁血红蛋白还原酶系统的组成部分）的结构特性。更具体地说，1) 为了解决在寄生适应过程中获得的 A. suum 细胞色素 b_5 的结构特性， 2）通过在线虫中表达前体细胞色素b_5来分析其生物合成模式，即A.suum细胞色素b_5的前序列的作用。成就1）以1.8A分辨率解析了A.suum细胞色素b_5的晶体结构。展示了与需氧哺乳动物细胞色素b_5（红细胞型，可溶性）不同的结构。 A. suum 血红蛋白和细胞色素 b_5 强烈表明它们是生理反应伙伴。免疫组织化学和免疫印迹研究表明，A. suum 细胞色素 b_5 位于皮下组织和肠周液中，并且细胞色素是一种秘书蛋白。提出了一个工作假设，即细胞色素 b_5 是一种专门适应宿主肠腔低氧张力的蛋白质。为了检验这一假设，细胞色素 b_5 物种被研究。对秀丽隐杆线虫基因数据库进行调查，其中已完成基因组计划，发现了四种秀丽隐杆线虫细胞色素_b5，尽管它们均不具有前序列，对这四个物种的亲水分析表明其中两种是可溶的。蛋白质，并且两者之一与 A. suum 细胞色素 b_5 表现出最高的同源性，但是，两者都没有被检测为表达序列标签 2) 由于效率低，目前正在检查实验条件。 3)使用Biacore进行亲和力分析，采用猪蛔虫和人细胞色素b_5作为配体，表明后者与肠周血红蛋白的亲和力低于前者。这些结果支持了上述工作假设。

项目成果

期刊论文数量（0）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Unique structure of Ascaris suum b_5-type cytochrome : an additional a-helix and positively charged residues on the surface domain interact with redox partners

猪蛔虫 b_5 型细胞色素的独特结构：额外的 a 螺旋和表面结构域上的带正电残基与氧化还原伙伴相互作用

DOI：
发表时间：
2006
期刊：
Biochemical Journal 394(2)
影响因子：
0
作者：
Yokota;T.;Nakajima;Y.;Yamakura;F.;Sugio;S.;Hashimoto;M.;Takamiya S
通讯作者：
Takamiya S

Ascaris suum cytochrome b_5,an indispensable component for reduction of oxygen-avid ferric hemoglobin and antioxidation

猪蛔虫细胞色素b_5是还原嗜氧铁血红蛋白和抗氧化不可缺少的成分

DOI：
发表时间：
2007
期刊：
影响因子：
0
作者：
Yokota;T.;Nakajima;Y.;Yamakura;F.;Sugio;S.;Hashimoto;M.;Takamiya S;Takehiro Yokota;Shinzaburo Takamiya;Shinzaburo Takamiya
通讯作者：
Shinzaburo Takamiya

Ascaris suum cytochrome 65, an adult-specific secretory protein reducing oxygen-avid ferric hemoglobin

猪蛔虫细胞色素 65，一种成人特异性分泌蛋白，可还原嗜氧铁血红蛋白