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Obesity and oxidative stress: involvement of SOD in the physiology of metabolic syndrome and atherosclerosis

肥胖和氧化应激：SOD 参与代谢综合征和动脉粥样硬化的生理学

基本信息

批准号：
22590563
负责人：
OOKAWARA Tomomi
金额：
$ 2.91万
依托单位：
Hyogo University of Health Sciences
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2010
资助国家：
日本
起止时间：
2010 至 2012
项目状态：
已结题

项目摘要

Recombinant extracellular superoxide dismutases were purified and the heparin affinity was investigated. 104 micrograms of the wild-type human enzyme was purified from 473mL of cultured medium of recombinant EC-SOD expressing cell line HECY6-7 through three steps of chromatography, heparin-affinity, hydroxyapatite, and gel-filtration. Removing the N-glycan reduced reactivity of ConA, and increased both of specific activity and heparin affinity of the enzyme. Two mutation studies at heparin binding domain (substitution of amino acid, Glu237Lys and doubling basic amino acid cluster, KERKKRRRESEC => KERKKRRRESECKEKKRRRESEC) gave rise to higher heparin binding EC-SOD. Another two mutation that moved the position of N-glycan (N121Q/Q207N and N121Q/S234N) affected the heparin affinity nature of the enzyme. These results suggest that the structure and position of N-glycan play an important role in the function and localization of EC-SOD.

纯化重组细胞外超氧化物歧化酶，并研究肝素亲和力。通过层析、肝素亲和、羟基磷灰石和凝胶过滤三个步骤，从473 mL表达重组EC-SOD的细胞系HECY 6 -7的培养基中纯化104微克野生型人酶。去除N-聚糖降低了ConA的反应活性，提高了酶的比活力和肝素亲和力。肝素结合域的两个突变研究（氨基酸取代，Glu 237 Lys和加倍碱性氨基酸簇，KERKKRRRESEC => KERKKRRRESECKEKKRRRESEC）产生了更高的肝素结合EC-SOD。移动N-聚糖位置的另外两个突变（N121 Q/Q207 N和N121 Q/S234 N）影响酶的肝素亲和力性质。这些结果表明，N-聚糖的结构和位置对EC-SOD的功能和定位起着重要的作用。

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）

N-Glycosylation profiling of recombinant mouse extracellular superoxide dismutase produced in Chinese hamster ovary cells

中国仓鼠卵巢细胞产生的重组小鼠胞外超氧化物歧化酶的 N-糖基化分析

DOI：
10.1007/s10719-011-9333-6
发表时间：
2011
期刊：
Glycoconj J
影响因子：
0
作者：
Okamoto I;Arao T;Miyazaki M;Satoh T;Okamoto K;Tsunoda T;Nishio K;Nakagawa K.;Korekane H
通讯作者：
Korekane H

Structural switching of Cu,Zn-superoxide dismutases at loop VI: insights from the crystal structure of 2-mercaptoethanol-modified enzyme.

Cu，Zn-塞氧化物歧化酶在环VI上的结构切换：从2-羟基乙醇改性酶的晶体结构中的见解。