高等学校化学学报

高等学校化学学报 ›› 2002, Vol. 23 ›› Issue (6): 1086.

• 研究论文 • 上一篇    下一篇

兔肝金属硫蛋白亚型异构体的高效液相色谱分离与ESI-MS,MALDI-TOF-MS鉴定

赵锐1, 纪建国1, 袁洪生1, 管华诗2, 茹炳根1   

  1. 1. 蛋白质组学研究室, 蛋白质工程及植物基因工程国家重点实验室, 北京大学生命科学院, 北京 100871;
    2. 青岛海洋大学海洋药物与食品研究所, 青岛 266003
  • 收稿日期:2001-03-22 出版日期:2002-06-24 发布日期:2002-06-24
  • 通讯作者: 茹炳根(1963年出生),男,教授,博士生导师,从事蛋白质化学与蛋白质组学研究.E-mail:rulab@pku.edu.cn E-mail:rulab@pku.edu.cn

Separation of Rabbit Liver Metallothionein Sub-isoforms by RP-HPLC with MALDI-TOF-MS Detection

ZHAO Rui1, JI Jian-Guo1, YUAN Hong-Sheng1, GUAN Hua-Shi2, RU Bing-Gen1   

  1. 1. Proteomics Research Group, National Laboratory of Protein Engineering & Plant Genetic Eengineering, Peking University, Beijing 100871,China;
    2. Institute of Marine Drugs and Foods, Ocean University of Qingdao, Qingdao 266003, China
  • Received:2001-03-22 Online:2002-06-24 Published:2002-06-24

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被引次数

17

摘要： 由于金属硫蛋白(MT)基因的多态性,决定不同亚型的MT异构体的存在,MT亚型异构体的结构是MT功能研究的基础.通过离子交换柱可将MT分成MT-1和MT-2两个异构体,用不同条件的反相高效液相色谱(RP-HPLC)可将MT-1和MT-2分成不同的亚型异构体,并利用MALDI-TOFMS和LC-ESI-MS对比确定了它们的分子量.结果表明,兔肝MT在不同的pH条件下分离得到不同分子量的亚型异构体.在酸性条件下,MT-1可分为2个主要亚型异构体,分子量分别为6149.0和6244.5,而MT-2主要分为3个亚型异构体,分子量分别为6149.0,6244.0和6127.0.MT-1和MT-2有2个亚型异构体分子量相同的异构体存在.在酸性条件下,MT-1的2个异构体及MT-2分子量为6127的亚型异构体可稳定存在.

关键词: 金属硫蛋白, 亚型异构体, RP-HPLC, 电喷雾离子化质谱, 基质辅助激光解析飞行时间质谱

Abstract: In order to study the sub-isoforms of MTand get their exact primary structure, the mixture and the individuals MT-1 and MT-2 of rabbit liver MTwere separated by different HPLCmethods such as anion exchange and RP-HPLCunder different conditions. The two main MTisoforms MT-1 and MT-2 could be easily isolated from the mixture of MTby anion exchange HPLC. The isoforms of MTs were further isolated by C18 RP-HPLCunder neutral and acidic conditions. All of the isolated isoforms and sub-isoforms were analyzed by MALDI-TOF-MS. The separation of MTisoforms can be obtained by anion exchange chromatography with 50 mmol/L Na2HPO4 NaCl. The sub-isoforms of rabbit liver MTcould be isolated by C18 RP-HPLC. The rabbit liver MT-1 has 3 main sub-isoforms and MT-2 has one sub-isoform. The experiment verified that under different conditions the sub-isoforms of rabbit liver MThas a different apparent molecular mass. This experiment predicted that there would exist a stable peptide structure of MT-2 or its fragment. The primary structure of this peptide component was also studied.

Key words: Metallothionein, Sub isoform, RP-HPLC, ESI-MS, MALDI-TOF-MS

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