海南产T-超家族芋螺毒素B8.1T合成及活性分析

生物技术通报 ›› 2013, Vol. 0 ›› Issue (7): 184-188.

海南产T-超家族芋螺毒素B8.1T合成及活性分析

吴勇,吴潇洒,长孙东亭,罗素兰

（海南大学热带生物资源教育部重点实验室海口市海洋药物重点实验室，海口 570228）

收稿日期:2013-01-16 修回日期:2013-07-19 出版日期:2013-07-19 发布日期:2013-09-02
作者简介:吴勇，男，助理研究员，研究方向：海洋药物与生物技术；E-mail：wys211@163.com
基金资助:
国家国际科技合作专项（2011DFR31210），国家自然科学基金项目（81160503），长江学者和创新团队发展计划（IRT1123），海南大学青年基金项目（qnjj1010）

Synthesis and Bioactivity of T-superfamily Conotoxin B8.1T Native to Hainan

Wu Yong, Wu Xiaosa, Zhangsun Dongting, Luo Sulan

（Key Laboratory of Tropical Biological Resources of Ministry of Education，Hainan University；

Received:2013-01-16 Revised:2013-07-19 Published:2013-07-19 Online:2013-09-02

摘要/Abstract

摘要： 从海南产桶形芋螺中发现了一个新的T-超家族芋螺毒素B8.1T，其氨基酸序列为DCCPESPPCCH，具有两对二硫键，其连接方式为Cys2-Cys9，Cys3-Cys10。首先采用N-9-芴甲氧羰基（Fmoc）固相合成法合成了B8.1T线性肽，根据采用半胱氨酸保护基团的不同，探索了3种不同的氧化方法：（1）两步氧化法；（2）One-pot方法；（3）自由氧化法。同时对提高芋螺毒素氧化折叠效率的条件进行了优化，对B8.1T的生物活性进行了初步的研究。结果表明，3种方法都可合成正确折叠的芋螺毒素B8.1T，其中两步氧化法和One-pot法合成效率高于自由氧化法，自由氧化折叠条件的优化表明，折叠反应添加物等能提高正确折叠产物的积累效率。B8.1T的动物活性试验显示，当对小鼠进行颅内给药时，对其行为活动产生抑制效应，而对肌肉给药的金鱼无明显影响。

关键词: T-超家族芋螺毒素, 固相多肽合成, 氧化折叠, 二硫键形成

Abstract: A new T-superfamily conotoxin B8.1T with sequence DCCPESPPCCH was discovered from C. betulinus native to Hainan. Conotoxin B8.1T contains two disulphide bridges between Cys3-Cys9, Cys4-Cys10. The linear conotoxin B8.1T was synthesized by Fmoc solid-phase synthesis method at first. Three oxidative methods were used for folding linear peptides according to different cysteine thiol protecting groups：（1）two-steps oxidative folding；（2）one-pot folding；（3）random one-step oxidative folding. Oxidative folding parameters were optimized to improve folding efficiency. The bioactivity of conotoxin B8.1T was assayed. The results demonstrated that all the three strategies can be used to produce native conotoxin B8.1T. However, the efficiencies of the two-step and one-pot methods were higher than that of the one-step. The presence of folding additives helped to improve native isomer accumulating by parameters’ optimization. The bioassay showed that conotoxin B8.1T inhibited the movement behavior of mice by intracranial injections, while there was no effects on goldfish by intramuscular injections. The results would lay a solid foundation for follow-up synthesis and research of T-superfamily contoxins.

Key words: T-superfamily conotoxin, Solid peptide synthesis, Oxidative folding, Disulfide bond formation

吴勇,吴潇洒,长孙东亭,罗素兰. 海南产T-超家族芋螺毒素B8.1T合成及活性分析[J]. 生物技术通报, 2013, 0(7): 184-188.

Wu Yong, Wu Xiaosa, Zhangsun Dongting, Luo Sulan. Synthesis and Bioactivity of T-superfamily Conotoxin B8.1T Native to Hainan[J]. Biotechnology Bulletin, 2013, 0(7): 184-188.

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