Department of Chemistry and Pharmacy, Chair of Food Chemistry, Emil Fischer Center, University of Erlangen-Nuremberg, Erlangen, Germany.
J Agric Food Chem. 2013 Jul 17;61(28):6971-81. doi: 10.1021/jf401549j. Epub 2013 Jul 3.
Nonenzymatic post-translational protein modifications (nePTMs) result in changes of the protein structure that may severely influence physiological and technological protein functions. In the present study, ultrahigh-performance liquid chromatography-electrospray ionization tandem mass spectrometry (UHPLC-ESI-MS/MS) was applied for the systematic identification and site-specific analysis of nePTMs of β-lactoglobulin in processed milk. For this purpose, β-lactoglobulin, which had been heated with lactose under conditions to force nePTM formation (7 d/60 °C), was screened for predicted modifications by using full scans and enhanced resolution scan experiments combined with enhanced product ion scans. Thus, the main glycation, glycoxidation, oxidation, and deamidation products of lysine, arginine, methionine, cysteine, tryptophan, and asparagine, as well as the N-terminus, were identified. Using these MS data, a very sensitive scheduled multiple reaction monitoring method suitable for the analysis of milk products was developed. Consequently, 14 different PTM structures on 25 binding sites of β-lactoglobulin were detected in different milk products.
非酶翻译后蛋白质修饰（nePTMs）会导致蛋白质结构发生变化，从而严重影响生理和技术蛋白功能。在本研究中，采用超高效液相色谱-电喷雾串联质谱（UHPLC-ESI-MS/MS）系统鉴定了加工牛奶中β-乳球蛋白的 nePTMs，并对其进行了位点特异性分析。为此，使用全扫描和增强分辨率扫描实验结合增强产物离子扫描，对在热诱导 nePTM 形成条件下（7d/60°C）与乳糖共热的β-乳球蛋白进行了预测修饰的筛选。因此，鉴定出赖氨酸、精氨酸、蛋氨酸、半胱氨酸、色氨酸和天冬酰胺的主要糖化、糖基化氧化和脱酰胺产物，以及 N 端。利用这些 MS 数据，开发了一种非常灵敏的适用于乳制品分析的预定多重反应监测方法。结果在不同的乳制品中检测到 25 个结合位点上的 14 种不同的 PTM 结构。
