Formylation of His Residues at Position 2 in Peptide b Ions due to Noncanonical Cα-CO Bond Cleavage: Assignment of the m/z 166 Product Ion in Proteome Data

IF 1.7 3区化学 Q4 BIOCHEMICAL RESEARCH METHODS

Rapid Communications in Mass Spectrometry Pub Date : 2025-10-23 DOI:10.1002/rcm.10154

Sanjeev Kumar, M. Vijayasarathy, M. A. Venkatesha, P. Balaram

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引用次数: 0

Abstract

Rationale

The low m/z region (less than 300) in the MS/MS spectra of peptides is rich with product ions arising from neutral losses and noncanonical bond cleavages, resulting from the gas-phase chemistry of low mass b_n/y_n ions containing reactive side chains. This study explores the ion chemistry arising from the presence of His residues at the N-terminus of peptides, focusing on the product ion at m/z 166 often designated as His-related ion.

Methods

Tandem mass spectrometry analysis of model His containing synthetic di and tripeptides was carried out on an ion-trap mass spectrometer. Isotope labelling (¹³C) of the residue(1) CO group permitted monitoring the fate of the labelled carbon atom. A dataset of tryptic peptides containing the m/z 166 ion in their MS/MS spectra was extracted from the consensus human-HCD proteome data available from the NIST database.

Results

MS/MS analysis of the tripeptide Ala-His-Ala-amide (AHA*), together with isotopically labelled analogs N^α-formyl histidyl ion (fH) amide and synthetic N^α-formyl His amide establishes that the product ion at m/z 166.061 corresponds to the protonated form of the N^α-formyl histidyl ion. Studies with¹³C labelled LH amide confirm fH arises by C^α-CO bond cleavage in the XH b₂ ion. Proteomic data analysis confirms that this unusual cleavage is widely observed in tryptic peptides. Examination of the top 500 peptides based on intensity of m/z 166 revealed that the overwhelming majority contain N-terminus XH motifs. Examinations of the top 100 sequences from a dataset of 6064 peptides lacking His that yielded m/z 166 revealed that the majority of 85 sequences arise from AQ b₂ ions by neutral loss of 2NH₃.

Conclusions

In peptide sequences containing N-terminus XH motifs, the histidine imidazole side chain facilitates C^α-CO bond cleavage at residue X.

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非规范Cα-CO键断裂导致肽b离子2位His残基甲酰化：蛋白质组数据中m/ z166产物离子的分配

多肽的MS/MS谱中的低m/z区（小于300）富含由中性损失和非规范键断裂引起的产物离子，这是由含有活性侧链的低质量bn/yn离子的气相化学反应引起的。本研究探讨了由多肽n端存在的His残基引起的离子化学，重点研究了位于m/z 166的产物离子，通常被称为His相关离子。方法采用离子阱质谱仪对含合成二肽和三肽的His模型进行串联质谱分析。残留物(1)CO基团的同位素标记（13C）允许监测标记的碳原子的命运。从NIST数据库中获得的一致的人类- hcd蛋白质组数据中提取了MS/MS光谱中含有m/z 166离子的色氨酸肽数据集。结果对三肽α- His- ala -amide （AHA*）、同位素标记的类似物n - α-甲酰基组氨酸（fH）酰胺和合成的n - α-甲酰基组氨酸酰胺进行MS/MS分析，产物离子在m/z 166.061处对应于n - α-甲酰基组氨酸离子的质子化形式。用13c标记LH酰胺的研究证实，fH是由xhb2离子中的c - α- co键断裂引起的。蛋白质组学数据分析证实，这种不寻常的切割在色氨酸肽中广泛观察到。基于m/z 166强度的前500个多肽检测显示，绝大多数含有n端XH基序。从6064个缺乏His的肽集中检测前100个序列，得到m/z 166，结果表明85个序列中的大多数来自AQ b2离子，由2NH3的中性损失引起。结论在含有n端XH基序的肽序列中，组氨酸咪唑侧链有利于残基X处c - α- co键的断裂。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Rapid Communications in Mass Spectrometry 化学-分析化学

CiteScore

4.10

自引率

5.00%

发文量

219

审稿时长

2.6 months

期刊介绍： Rapid Communications in Mass Spectrometry is a journal whose aim is the rapid publication of original research results and ideas on all aspects of the science of gas-phase ions; it covers all the associated scientific disciplines. There is no formal limit on paper length ("rapid" is not synonymous with "brief"), but papers should be of a length that is commensurate with the importance and complexity of the results being reported. Contributions may be theoretical or practical in nature; they may deal with methods, techniques and applications, or with the interpretation of results; they may cover any area in science that depends directly on measurements made upon gaseous ions or that is associated with such measurements.