药学学报, 2015, 50(7): 910-915
引用本文:
王庆华, 梁兰, 陈晶晶, 巩婷, 侯琦, 杨金玲, 朱平. 毕赤酵母Muts与Mut+重组子表达蝎毒镇痛活性肽BmK AngM1水平比较[J]. 药学学报, 2015, 50(7): 910-915.
WANG Qing-hua, LIANG Lan, CHEN Jing-jing, GONG Ting, HOU Qi, YANG Jin-ling, ZHU Ping. The expression of BmK AngM1 in Muts and Mut+ recombinants of Pichia pastoris[J]. Acta Pharmaceutica Sinica, 2015, 50(7): 910-915.

毕赤酵母Muts与Mut+重组子表达蝎毒镇痛活性肽BmK AngM1水平比较
王庆华, 梁兰, 陈晶晶, 巩婷, 侯琦, 杨金玲, 朱平
中国医学科学院、北京协和医学院药物研究所, 天然药物活性物质与功能国家重点实验室 & 卫生部天然药物生物合成重点实验室, 北京 100050
摘要:
蝎毒镇痛活性肽BmK AngM1是从东亚钳蝎 (Buthus martensii Karsch) 蝎毒中分离得到的一种新型长链蝎毒素, 其镇痛活性强且毒性低, 有望开发成镇痛新药。本文将BmK AngM1基因转入毕赤酵母 (Pichia pastoris) GS115, 筛选得到甲醇利用缓慢型 (Muts) 和快速型 (Mut+) 的重组子; 采用实时荧光定量PCR方法, 检测了Mut+重组子中BmK AngM1基因的拷贝数, 筛选出含单拷贝BmK AngM1基因的Mut+重组子; 在相同培养条件下, 比较了含单拷贝BmK AngM1基因的Muts和Mut+重组子表达BmK AngM1的水平。结果表明, Muts重组子中BmK AngM1基因转录水平是Mut+重组子的2.7倍, Muts重组子中BmK AngM1蛋白表达量是Mut+重组子的1.5倍。因此, Muts重组子比Mut+重组子具有更强的BmK AngM1表达能力。
关键词:    蝎毒镇痛活性肽      毕赤酵母      基因拷贝数      Muts重组子      Mut+重组子     
The expression of BmK AngM1 in Muts and Mut+ recombinants of Pichia pastoris
WANG Qing-hua, LIANG Lan, CHEN Jing-jing, GONG Ting, HOU Qi, YANG Jin-ling, ZHU Ping
State Key Laboratory of Bioactive Substance and Function of Natural Medicines and Key Laboratory of Biosynthesis of Natural Products, Ministry of Health of PRC, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China
Abstract:
BmK AngM1 is a long-chain scorpion toxin purified from the venom of Buthus martensii Karsch. It has been reported to exhibit evident analgesic effect and low toxicity, and has the potential to be a novel analgesic drug. The BmK AngM1 gene was transformed into Pichia pastoris GS115. Mut+ and Muts recombinant strains were screened by phenotype and Mut+ recombinant strains were used to detect BmK AngM1 gene copy number in the real-time PCR. Expression of BmK AngM1 in the Mut+ recombinant strain was compared with that of the Muts recombinant strain with the same single copy of BmK AngM1 gene under the same condition. The results indicated that the transcription level of BmK AngM1 gene in the Muts recombinant strain was 2.7 fold of that in the Mut+ recombinant strain in the real-time PCR, and the expression of BmK AngM1 in the Muts recombinant strain was 1.5 fold of that in the Mut+ recombinant strain. Therefore, Muts recombinant strain showed better ability to express BmK AngM1 than Mut+ recombinant strain.
Key words:    BmK AngM1    Pichia pastoris    gene copy number    Muts recombinant    Mut+ recombinant   
收稿日期: 2015-04-29
基金项目: 国家自然科学基金资助项目 (30873384); 北京市自然科学基金资助项目 (7122115); 天然药物活性物质与功能国家重点实验室自主课题.
通讯作者: 杨金玲, 朱平
Email: yangjl@imm.ac.cn;zhuping@imm.ac.cn
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