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  

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

作　　者：储艳秋[1] 戴兆云[2] 蒋公羽[1] 郑松柏[2] 陈玺[2] 丁传凡[1] 

机构地区：[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

出　　处：《高等学校化学学报》2009年第5期869-875,共7页Chemical Journal of Chinese Universities

基　　金：国家自然科学基金(批准号:25027004)资助

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5×10-5～3.00×10-4mol/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.To explore the non-covalent interaction between reduced tripeptide glutathione and L-aromatic amino acids, a stoiehiometry 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 eleetrospray 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 MS^2 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.

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 

分 类 号：O657.63[理学—分析化学] O641.4[理学—化学]