Hefei National Laboratory for Physical Sciences at the Microscale, School of Chemistry and Materials Science, University of Science and Technology of China, 230026, Hefei, China.
Hefei National Laboratory for Physical Sciences at the Microscale, School of Chemistry and Materials Science, University of Science and Technology of China, 230026, Hefei, China; National Synchrotron Radiation Laboratory, University of Science and Technology of China, Hefei, Anhui, 230029, PR China.
Anal Chim Acta. 2021 Jan 2;1141:120-126. doi: 10.1016/j.aca.2020.10.036. Epub 2020 Oct 22.
Investigation of protein-ligand interactions in physiological conditions is crucial for better understanding of biochemistry because the binding stoichiometry and conformations of complexes in biological processes, such as various types of regulation and transportation, could reveal key pathways in organisms. Nanoelectrospray ionization mass spectrometry is widely used in studies of biological processes and systems biology. However, non-volatile salts in biological fluid may adversely interfere with nanoelectrospray ionization mass spectrometry. In this study, the previously developed method of induced nanoelectrospray ionization was used to facilitate in situ desalting of protein in solutions with high concentrations of non-volatile salts, and direct investigation of protein-ligand interactions for the first time. In situ desalting occurred at the tip of emitters within a short period lasting for a few to tens of milliseconds, enabling the maintenance of nativelike conditions compatible with mass spectrometry measurements. Induced nanoelectrospray ionization was driven by pulsed potential and exhibited microelectrophoresis effect in each spray cycle, which is not observed in conventional nanoelectrospray ionization because the continuous spray procedure is driven by direct current. Microelectrophoresis caused desalting through micron-sized spray emitters (1-20 μm), as confirmed experimentally with proteins in 100 mM NaCl solution. The method developed in this study has been further illustrated as a potential option for fast and direct identification of protein-ligand (small molecules or metal ions) interactions in complex samples. The results of this study demonstrate that the newly developed method may represent a reliable approach for investigations of proteins and protein complexes in biological samples.
研究生理条件下的蛋白质-配体相互作用对于更好地理解生物化学至关重要，因为在各种类型的调节和运输等生物过程中，复合物的结合计量和构象可以揭示生物体中的关键途径。纳喷雾电离质谱广泛应用于生物过程和系统生物学的研究。然而，生物流体中的非挥发性盐可能会对纳喷雾电离质谱产生不利干扰。在本研究中，我们使用先前开发的诱导纳喷雾电离方法，促进高浓度非挥发性盐溶液中蛋白质的原位脱盐，并首次直接研究蛋白质-配体相互作用。原位脱盐发生在短时间内（持续几到几十毫秒）的发射器尖端，从而维持与质谱测量兼容的天然条件。诱导纳喷雾电离由脉冲电势驱动，并在每个喷雾循环中表现出微电泳效应，这在传统的纳喷雾电离中观察不到，因为连续的喷雾过程是由直流电驱动的。微电泳通过微米级的喷雾发射器（1-20 μm）进行脱盐，这在用 100 mM NaCl 溶液中的蛋白质进行的实验中得到了证实。本研究中开发的方法进一步说明了它是一种快速、直接识别复杂样品中蛋白质-配体（小分子或金属离子）相互作用的潜在选择。本研究的结果表明，新开发的方法可能代表了一种可靠的方法，用于研究生物样品中的蛋白质和蛋白质复合物。
