高等学校化学学报 ›› 2009, Vol. 30 ›› Issue (5): 869.

• 研究论文 • 上一篇    下一篇

电喷雾质谱法研究谷胱甘肽与L型芳香性氨基酸非共价复合物

储艳秋1, 戴兆云2, 蒋公羽1, 郑松柏2, 陈玺2, 丁传凡1   

    1. 复旦大学化学系激光化学研究所, 上海 200433;
    2. 复旦大学附属华东医院, 上海 200040
  • 收稿日期:2009-01-15 出版日期:2009-05-10 发布日期:2009-05-10
  • 通讯作者: 储艳秋, 男, 博士, 副教授, 主要从事有机分子和生物大分子质谱研究, E-mail: chuyq@fudan.edu.cn; 丁传凡, 男, 博士, 教授, 主要从事生物质谱研究, E-mail: cfding@fudan.edu.cn
  • 基金资助:

    国家自然科学基金(批准号: 25027004)资助.

Studies on Non-covalent Complexes of Glutathione with L-Aromatic Amino Acids by Electrospray Ionization Mass Spectrometry

CHU Yan-Qiu1* , DAI Zhao-Yun2, JIANG Gong-Yu1, ZHENG Song-Bai2, CHEN Xi2, DING Chuan-Fan1*   

    1. Laser Chemistry Institute, Chemistry Department, Fudan University, Shanghai 200433, China;
    2. Huadong Hospital Affiliated with Fudan University, Shanghai 200040, China
  • Received:2009-01-15 Online:2009-05-10 Published:2009-05-10
  • Contact: CHU Yan-Qiu, E-mail: chuyq@fudan.edu.cn; DING Chuan-Fan, E-mail: cfding@fudan.edu.cn

摘要:

为探索谷胱甘肽和L型芳香性氨基酸的非共价相互作用, 将一定化学剂量比的还原型γ-谷胱甘肽分别与L型芳香性氨基酸(包括苯丙氨酸、酪氨酸和色氨酸)在室温和生理pH条件下混合后, 温育1 h, 生成非共价复合物, 并使反应完全. 电喷雾质谱测量结果揭示谷胱甘肽和L型芳香性氨基酸反应可以生成非共价复合物. 在二级串级质谱MS2测得的复合物碎片离子峰中, 除芳香性氨基酸离子峰外, 还包括谷胱甘肽及其它再次碎裂产生的b2和y2碎片离子, 进一步确认了非共价复合物的形成. 紫外光谱也证实了电喷雾质谱的实验结果. 为避免严重的离子化效率差异和质谱信号的相互抑制作用, 定量评估了谷胱甘肽和酪氨酸的相互作用, 结果显示反应物的初始浓度应该选择在5×10-5~3.00×10-4 mol/L范围内. 用质谱滴定法测定了谷胱甘肽与3个芳香性氨基酸非共价复合物的解离常数, 结果表明, 谷胱甘肽复合物的稳定性按Tyr, Trp和Phe次序依次增大.

关键词: L型芳香性氨基酸, 谷胱甘肽, 非共价复合物, 质谱滴定法, 解离常数

Abstract:

To explore the non-covalent interaction between reduced tripeptide glutathione and L-aromatic amino acids, a stoichiometry of reduced γ-glutathione and three L-aromatic amino acids, including phenylalanine, tyrosine and tryptophan were mixed respectively, and then incubated at room temperature and physiological pH conditions for 1 h to reach the equilibrium. The electrospray ionization mass spectrometry(ESI-MS) results indicate that glutathione and three L-aromatic amino acids could form non-covalent complexes, respectively. The primary fragment ions of complexes obtained from MS2 in a tandem mass spectrometer included aromatic amino acid, glutathione as well as its y2, b2, which confirmed the results of ESI-MS. Moreover, in UV spectroscopies, the discrepancy between complexes and reactants also confirmed the formation of complexes in aqueous solution. To avoid distinct ionization efficiency discrepancy and signal suppression in ESI-MS measurements, the interaction between glutathione and L-Tyrosine was evaluated quantitatively, which revealed that the reactant concentration should range from 5×10-5 to 3.00×10-4 mol/L. The dissociation constants of three complexes were determined using mass spectrometric titration method, the calculation results revealed that the stabilities of complexes formed by glutathione and three L-aromatic amino acids increased gradually according to the order of tyrosine, tryptophan and phenylalanine.

Key words: L-Aromatic amino acids, Glutathione, Non-covalent complexes, Mass spectrometric titration method, Dissociation constants

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