基于激光解析/离子化-飞行时间质谱技术的中药阿胶蛋白质组分析

Analysis of the traditional Chinese medicine Donkey-hide gelatin proteome based on surface enhanced laser desorption/ionization time-of-flight mass spectrometry

目的:应用激光解析/离子化-飞行时间质谱技术对传统中药阿胶蛋白/肽成分进行蛋白质组初步分析,建立阿胶蛋白质质量指纹图。方法:实验于2004-06/2005-03在湖南中医药大学蛋白质组学实验室完成。采用饱和硫酸胺沉淀,透析脱盐冻干法获得阿胶水溶性蛋白/肽,采用Ciphergen BiosystemsInc.ProteinChipRBiology System(美国PBSⅡ+)及配套的NP10芯片、激光解析/离子化-飞行时间质谱技术,分析阿胶Mr1500～13000区间蛋白质、肽分布及其相对分子质量。结果:Mr2000～4000区间显示4个相对分子质量峰,可能是2种肽;Mr4000～5000区间显示8个相对分子质量峰,可能为2种肽;Mr5000～5500区间显示16个相对分子质量峰,约为3种肽;Mr5500￣6200区间无峰;Mr8100～8200区间显示5个相对分子质量峰,可能为1种肽;Mr11200～11400区间显示5个相对分子质量峰,可能为1种蛋白质。不同质量浓度稳定获得的有意义蛋白/肽共计9个。不同相对分子质量肽之间差分别提示可能为丙氨酸、丝氨酸、苯丙氨酸残基相对分子质量。结论:通过分析,可以形成阿胶蛋白质/肽成分质量指纹图,可作为阿胶数字化质控标准;并为进一步分离、纯化及验证阿胶功能相关活性蛋白质/肽组分提供可靠信息。

AIM： To set up the proteomic fingerprinting of Donkey-hide gelatin by using surface enhanced laser desorption/ionization time-of-flight mass spectrometry （SELDI-TOF-MS）. METHODS： The experiment was performed at the Laboratory of Proteome, Hunan University of Traditional Chinese Medicine from June 2004 to March 2005. By means of dialysis-desalination-freeze drying with satisfying sulphuric acid amine sediment, the water soluble protein/ peptide from the Donkey-hide gelatin was gotten. Ciphergen Biosystems Inc.ProteinChip Biology System （America PBS Ⅱ ＋） and combined NP10array, SELDI-TOF-MS were adopted to analyze the Donkey-hide gelatin＇s Mr 1 500-13 000 Da protein, peptide distribution and its relative molecular mass （RMM）. RESULTS：There were 4 peaks, disl01aying 2 kinds of peptide between Mr 2 000-4 000 Da; 8 peaks, displaying 2 kinds of peptide between Mr 4 000-5 000 Da; 16 peaks, about 3 kinds of peptide between Mr 5 000-5 500 Da; no peak between Mr 5 500-6 200 Da; 5 peaks, belonged to a kind of peptide between Mr 8 100-8 200 Da; and 5 peaks, belonged to a kind of peptide between Mr 11 200-11 400 Da. There were 9 kinds of meaningful protein/peptide with different mass concentrations. It might display the lactamine, serine, phenyialanine residues from the disparities of different RMM. CONCLUSION： The Donkey-hide gelatin protein/peptide mass fingerprinting （PMF） can be obtained as digital quality control standard of the Donkey-hide gelatin. Reliable information on further isolation, purification and verification of the Donkey-hide gelatin protein/peptide is provided.