Department of Cardiology, The First Affiliated Hospital of USTC, Division of Life Sciences and Medicine, University of Science and Technology of China, Hefei, Anhui 230001, China.
School of Chemistry and Materials Science, University of Science and Technology of China, Hefei, Anhui 230001, China.
Anal Chem. 2022 Oct 11;94(40):13829-13833. doi: 10.1021/acs.analchem.2c02390. Epub 2022 Oct 2.
Native mass spectrometry, which takes a high concentration of ammonium acetate (NHOAc) for ionization, coupled with tedious and solvent-consuming purification, which separates proteins from complicated environments, has shown great potential for proteins and their complexes. A high level of nonvolatile salts in the endogenous intracellular environment results in serious ion suppression and has been one of the bottlenecks for native mass spectrometry, especially for protein complexes. Herein, an integrated protocol utilizing the inner surface of a micropipette for rapid purification, desorption, and ionization of peptide-metal interaction at subfemtomole level in cell lysate was demonstrated for native mass spectrometry. The methods showed robust and reproducibility in protein measurement within 1 min from various buffers. The cells expressing with various proteins were lysed and used to test our method. The specific interaction between the peptide-metal complex in cell lysates could be reserved and distinguished by mass spectrometry.
基于纳升电喷雾的原位质谱分析，由于采用高浓度的乙酸铵（NHOAc）进行离子化，以及繁琐且耗溶剂的蛋白质分离纯化步骤，其在蛋白质及其复合物的分析中显示出巨大的潜力。内源性细胞内环境中高水平的不可挥发盐会导致严重的离子抑制，这一直是原位质谱分析的瓶颈之一，尤其是对于蛋白质复合物。本文展示了一种整合方案，利用微吸管的内表面在细胞裂解液中实现肽-金属相互作用的快速纯化、解吸和离子化，用于原位质谱分析。该方法在 1 分钟内即可从各种缓冲液中进行蛋白质检测，表现出稳健性和重现性。表达不同蛋白质的细胞被裂解并用于测试我们的方法。通过质谱分析可以保留和区分细胞裂解液中肽-金属复合物的特异性相互作用。
