钠离子通道β4亚基糖基化的初步研究

doi:10.13523/j.cb.20140702

中国生物工程杂志

2014, Vol. 34

Issue (7): 10-16 DOI: 10.13523/j.cb.20140702

研究报告

钠离子通道β4亚基糖基化的初步研究

周婷婷^1,2, 潘传涌², 张建鹏², 金慧英¹

1. 南京军事医学研究所南京 210002;
2. 第二军医大学生物化学与分子生物学教研室上海 200433

The Research of the Glycosylation of Sodium Channel β4 Subunit

ZHOU Ting-ting^1,2, PAN Chuan-yong², ZHANG Jian-peng², JIN Hui-ying¹

1. East-China Institute for Medical Biotechniques, Nanjing 210002, China;
2. Department of Biochemistry & Molecular Biology, Second Military Medical University, Shanghai 200433, China

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摘要：

异常表达的糖蛋白与PD等多种神经退行性疾病有关。糖蛋白组学研究发现，电压门控钠离子通道β4亚基在PD病人脑组织中表达明显增加。为了深入探索β4亚基及其糖链在帕金森发生发展中的作用，采用PD转基因鼠对其表达进行验证，对其潜在的糖基化位点进行定点突变，构建重组表达质粒。结果发现，在新生PD转基因鼠和野生成鼠脑组织中有~38kDa蛋白条带表达，而在新生野生鼠脑组织中不表达；用PNGase F酶处理去除糖链后，~38kDa蛋白条带变成迁移速递更快的较小分子量条带，说明β4亚基是高度糖基化的蛋白，并且其糖基化与生长发育有关。将突变重组质粒转入HEK-293细胞和小鼠神经瘤细胞Neuro2A中表达，结果发现突变型质粒分子量明显低于野生型。为研究β4亚基及其糖链的功能提供了一定的实验数据并打下了基础。

关键词： 电压门控钠离子通道β4亚基; 糖基化; 帕金森病

Abstract:

Aberrant protein glycosylation plays major roles in neurodegenerative disease, including PD. Glycoproteomics showed that the glycosylation of sodium channel β4 was significantly increased in human brain tissue. β4-specific antibodies reacted in immunoblot assays with ~38 kDa band from the membrane fractions isolated from neonatal PD transgenic mice but not expressed in the neonatal wild-type mice. The molecular weight of ~38 kDa immunoreactive protein is in close agreement with previously reported, suggesting heavy glycosylation of this protein in adult wild-type and neonatal PD transgenic brain tissues. Enzymatic deglycosylation of the membrane preparations converted the 38 kDa band into a faster migrating protein, which was consistent with heavy glycosylation of this protein. The glycosylated state of β4 was developmentally regulated and was altered in disease state. We also expressed β4-wild type and deglycosylated mutant β4-MUT plasmids in HEK-293 and Neuro2A cells. These results lay a foundation of studying β4 subunit in the pathogenesis of PD. Further studies will focus on the effects of the oligosaccharides on the sodium channel activities and neuritic degeneration.

Key words: Sodium channel β4 subunit Glycosylation Parkinson’s disease

收稿日期: 2013-06-26 出版日期: 2014-07-25

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通讯作者: 金慧英 E-mail: jhying2013@126.com

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引用本文:

周婷婷, 潘传涌, 张建鹏, 金慧英. 钠离子通道β4亚基糖基化的初步研究[J]. 中国生物工程杂志, 2014, 34(7): 10-16.

ZHOU Ting-ting, PAN Chuan-yong, ZHANG Jian-peng, JIN Hui-ying. The Research of the Glycosylation of Sodium Channel β4 Subunit. China Biotechnology, 2014, 34(7): 10-16.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/10.13523/j.cb.20140702 或 https://manu60.magtech.com.cn/biotech/CN/Y2014/V34/I7/10

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