Institute of Chemical Technologies and Analytics, TU Wien (Vienna University of Technology), Getreidemarkt 9/164-IAC, A-1060, Vienna, Austria.
J Am Soc Mass Spectrom. 2017 Jan;28(1):77-86. doi: 10.1007/s13361-016-1483-0. Epub 2016 Sep 19.
In order to better understand biological events, lectin-glycoprotein interactions are of interest. The possibility to gather more information than the mere positive or negative response for interactions brought mass spectrometry into the center of many research fields. The presented work shows the potential of a nano-electrospray gas-phase electrophoretic mobility molecular analyzer (nES GEMMA) to detect weak, noncovalent, biospecific interactions besides still unbound glycoproteins and unreacted lectins without prior liquid phase separation. First results for Sambucus nigra agglutinin, concanavalin A, and wheat germ agglutinin and their retained noncovalent interactions with glycoproteins in the gas phase are presented. Electrophoretic mobility diameters (EMDs) were obtained by nES GEMMA for all interaction partners correlating very well with molecular masses determined by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) of the individual molecules. Moreover, EMDs measured for the lectin-glycoprotein complexes were in good accordance with theoretically calculated mass values. Special focus was laid on complex formation for different lectin concentrations and binding specificities to evaluate the method with respect to results obtained in the liquid phase. The latter was addressed by capillary electrophoresis on-a-chip (CE-on-a-chip). Of exceptional interest was the fact that the formed complexes could be sampled according to their size onto nitrocellulose membranes after gas-phase separation. Subsequent immunological investigation further proved that the collected complex actually retained its native structure throughout nES GEMMA analysis and sampling. Graphical Abstract ᅟ.
为了更好地理解生物事件，糖蛋白与凝集素的相互作用引起了人们的兴趣。与仅仅检测相互作用的阳性或阴性结果相比，质谱法有可能收集更多的信息，这使得它成为许多研究领域的中心。本工作展示了纳米电喷雾气相电泳迁移率分子分析器（nES GEMMA）的潜力，它可以在无需预先进行液相分离的情况下，检测到弱的、非共价的、生物特异性相互作用，以及未结合的糖蛋白和未反应的凝集素。首先介绍了黑接骨木凝集素、伴刀豆球蛋白 A 和麦胚凝集素及其在气相中与糖蛋白的保留非共价相互作用的结果。通过 nES GEMMA 获得了所有相互作用伙伴的电泳迁移率直径（EMD），与通过基质辅助激光解吸/电离质谱（MALDI-MS）对单个分子进行测定的分子量非常吻合。此外，测量的凝集素-糖蛋白复合物的 EMD 值与理论计算的质量值非常吻合。特别关注不同的凝集素浓度和结合特异性形成复合物，以评估该方法在液相中获得的结果。后者通过芯片毛细管电泳（CE-on-a-chip）来解决。特别有趣的是，形成的复合物可以在气相分离后根据其大小按照尺寸采样到硝酸纤维素膜上。随后的免疫学研究进一步证明，在 nES GEMMA 分析和采样过程中，收集到的复合物实际上保留了其天然结构。
