Pharmaceutical Biochemistry and Bioanalytics, Pharmaceutical Institute, University of Bonn, An der Immenburg 4, 53121 Bonn, Germany.
Anal Chem. 2020 Aug 18;92(16):10920-10924. doi: 10.1021/acs.analchem.0c02151. Epub 2020 Aug 6.
Disulfide bonds within cysteine-rich peptides are important for their stability and biological function. In this respect, the correct disulfide connectivity plays a decisive role. The differentiation of individual disulfide-bonded isomers by traditional high-performance liquid chromatography (HPLC) and mass spectrometry (MS) is limited due to the similarity in physicochemical properties of the isomers sharing the same amino acid sequence. By using trapped ion mobility spectrometry-mass spectrometry (TIMS-MS), several 2- and 3-disulfide-bonded isomers of the μ-conotoxin PIIIA were investigated for their distinguishability by collision cross section (CCS) values and their characteristic mobilogram traces. The isomers could be differentiated by TIMS-MS and also identified in mixing experiments. Thus, TIMS-MS provides a highly valuable and enriching addition to standard HPLC and MS analysis of conformational isomers of disulfide-rich peptides and proteins.
半胱氨酸丰富肽中的二硫键对于它们的稳定性和生物功能很重要。在这方面，正确的二硫键连接起着决定性的作用。由于具有相同氨基酸序列的异构体在物理化学性质上相似，因此传统的高效液相色谱 (HPLC) 和质谱 (MS) 对单个二硫键异构体的区分受到限制。通过使用被困离子淌度谱-质谱 (TIMS-MS)，研究了 μ-芋螺毒素 PIIIA 的几种 2-和 3-二硫键异构体的可区分性，方法是通过碰撞截面 (CCS) 值和它们的特征淌度图轨迹。TIMS-MS 可以区分异构体，并且也可以在混合实验中鉴定。因此，TIMS-MS 为富含二硫键的肽和蛋白质构象异构体的标准 HPLC 和 MS 分析提供了非常有价值和丰富的补充。
