Department of Chemistry, Pohang University of Science and Technology (POSTECH) , Pohang, 37673, South Korea.
Department of Chemistry, Korea University , Seoul, 02841, South Korea.
J Am Chem Soc. 2015 Dec 9;137(48):15322-9. doi: 10.1021/jacs.5b10648. Epub 2015 Nov 24.
Mass spectrometry (MS)-based analysis using enzymatic digestion is widely used for protein sequencing and characterization. The large number of peptides generated from proteolysis, however, suppresses the signal of peptides with low ionization efficiency, thus precluding their observation and analysis. This study describes a technique for improved analysis of peptic peptides by adding the synthetic receptor cucurbit[7]uril (CB[7]), which binds selectively to peptides with N-terminal aromatic residues. Capturing the N-terminal phenylalanine (Phe) of peptides using CB[7] enhances the peptide abundances both in electrospray ionization MS and in matrix-assisted laser desorption ionization MS. Moreover, collision-induced dissociation (CID) of the CB[7]·peptide complex ions generates b- and y-type fragment ions with higher sequence coverage than those generated with uncomplexed peptides. The signal enhancement mediated by CB[7] is attributed to an increase in the peptide proton affinities upon CB[7] complexation. The mechanistic details of the fragmentation process are discussed on the basis of the structures of the complex ions obtained from ion mobility (IM) measurements and molecular modeling. This study demonstrates a novel and powerful approach to the enhancement of protein and peptide analysis using a synthetic receptor, without the need for new instrumentation, chemical modifications, or specialized sample preparation. The simplicity and potential generality of this technique should provide a valuable asset in the toolbox of routine protein and peptide analysis.
基于酶解的质谱（MS）分析广泛用于蛋白质测序和表征。然而，从蛋白水解产生的大量肽会抑制电离效率低的肽的信号，从而阻止对其进行观察和分析。本研究描述了一种通过添加合成受体葫芦[7]脲（CB[7]）来改善肽酶解产物分析的技术，该受体可以选择性地与具有 N 端芳香族残基的肽结合。使用 CB[7]捕获肽的 N 端苯丙氨酸（Phe）可增强电喷雾电离 MS 和基质辅助激光解吸电离 MS 中肽的丰度。此外，CB[7]·肽配合物离子的碰撞诱导解离（CID）产生的 b-和 y-型片段离子比未与 CB[7]结合的肽产生的片段离子具有更高的序列覆盖率。CB[7]介导的信号增强归因于 CB[7]配合物形成后肽质子亲和力的增加。基于通过离子淌度（IM）测量和分子建模获得的配合物离子结构，讨论了碎裂过程的机理细节。本研究展示了一种使用合成受体增强蛋白质和肽分析的新颖而强大的方法，而无需新的仪器、化学修饰或专门的样品制备。该技术的简单性和潜在通用性应为常规蛋白质和肽分析工具包提供有价值的资产。
