Hunter A P, Games D E
Mass Spectrometry Research Unit, University College of Swansea, UK.
Rapid Commun Mass Spectrom. 1994 Jul;8(7):559-70. doi: 10.1002/rcm.1290080713.
The phosphorylation sites in a model phosphoprotein, alpha s1-casein from bovine milk, have been identified by tryptic peptide mapping (Gibson and Cohen, Methods Enzymol. vol. 193, p. 480 (1990)) employing reversed-phase high performance liquid chromatography (RPHPLC)/electrospray ionization mass spectrometry (ES-MS); by infusion tandem mass spectrometry (MS/MS) and LC/MS/MS in neutral loss mode of tryptic digests of alpha s1-casein, in which the characteristic neutral loss of phosphoric acid by phosphopeptides under collision-induced dissociation (CID) conditions is exploited to highlight phosphopeptides in a tryptic digest (Covey et al., in Methods in Protein Sequence Analysis, Jörnvall et al. (Eds), Birkhäuser Verlag, Basel 1991), and by a novel method, termed LC/CID-MS, in which phosphopeptides are located in mixtures of peptides by the generation and detection of phosphate-specific fragment ions during LC/ES-MS (Huddleston et al., J. Am. Soc. Mass Spectrom. vol. 4, p. 710 (1993)). An appraisal of the efficiency, sensitivity and practicality of each of these methods in the identification of phosphorylation sites in post-translationally modified proteins is given.
利用反相高效液相色谱（RPHPLC）/电喷雾电离质谱（ES-MS），通过胰蛋白酶肽图分析（吉布森和科恩，《酶学方法》第193卷，第480页（1990年）），已鉴定出一种模型磷蛋白——牛乳αs1-酪蛋白中的磷酸化位点；通过对αs1-酪蛋白胰蛋白酶消化产物进行中性丢失模式下的进样串联质谱（MS/MS）和液相色谱/串联质谱（LC/MS/MS），利用磷酸肽在碰撞诱导解离（CID）条件下磷酸的特征性中性丢失来突出胰蛋白酶消化产物中的磷酸肽（科维等人，载于《蛋白质序列分析方法》，约恩瓦尔等人（编），比尔克希尔出版社，巴塞尔，1991年），以及通过一种称为LC/CID-MS的新方法，即在液相色谱/电喷雾质谱（LC/ES-MS）过程中通过生成和检测磷酸盐特异性碎片离子来确定肽混合物中的磷酸肽（赫德尔斯顿等人，《美国质谱学会杂志》第4卷，第710页（1993年））。本文对这些方法在鉴定翻译后修饰蛋白质磷酸化位点方面的效率、灵敏度和实用性进行了评估。
