Analysis of Glutamine Deamidation: Products, Pathways, and Kinetics.
Analysis of Glutamine Deamidation: Products, Pathways, and Kinetics.
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DOI：
10.1021/acs.analchem.9b03127
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发表时间：
2019-10-15
影响因子：
7.4
通讯作者：
Julian RR
中科院分区：
文献类型：
--
作者：
Riggs DL;Silzel JW;Lyon YA;Kang AS;Julian RR
自发的化学修饰在分子水平上对人类疾病和衰老起着重要作用。已知脱酰胺和异构化是长寿命人类蛋白质中最普遍的化学修饰之一，并且与越来越多的人类病理学有关，但是与这些过程相关的相对较小的化学变化提出了长期的分析挑战。尽管采用高分辨率质谱法极大地帮助了蛋白质组学研究中脱酰胺位点的鉴定，但异构化（和脱酰胺的异构体产物）的表征仍然极具挑战性。在此，我们提出了一种基于液相色谱/质谱的方法，用于快速表征的异构体产品的谷氨酰胺脱酰胺使用诊断片段，大量生产，并能够明确识别Glu和isoGlu。重要的是，信息碎片离子是通过正交碎片化途径产生的，从而能够同时检测两种异构形式，同时保持与鸟枪蛋白质组学的兼容性。此外，与isoGlu相关的诊断片段可精确定位修饰残基的位置。该技术的实用性是通过表征一系列含谷氨酰胺的肽在体外老化过程中产生的异构体产物来证明的。序列依赖性的产品配置文件，并检查脱酰胺连接的外消旋丰度。最后，比较谷氨酰胺脱酰胺，这是相对知之甚少，天冬酰胺脱酰胺已被更彻底地研究。
Spontaneous chemical modifications play an important role in human disease and aging at the molecular level. Deamidation and isomerization are known to be among the most prevalent chemical modifications in long-lived human proteins and are implicated in a growing list of human pathologies, but the relatively minor chemical change associated with these processes has presented a longstanding analytical challenge. Although the adoption of high-resolution mass spectrometry has greatly aided the identification of deamidation sites in proteomic studies, isomerization (and the isomeric products of deamidation) remain exceptionally challenging to characterize. Herein, we present a liquid chromatography/mass spectrometry-based approach for rapidly characterizing the isomeric products of Gln deamidation using diagnostic fragments that are abundantly produced and capable of unambiguously identifying both Glu and isoGlu. Importantly, the informative fragment ions are produced through orthogonal fragmentation pathways, thereby enabling the simultaneous detection of both isomeric forms while retaining compatibility with shotgun proteomics. Furthermore, the diagnostic fragments associated with isoGlu pinpoint the location of the modified residue. The utility of this technique is demonstrated by characterizing the isomeric products generated during in vitro aging of a series of glutamine-containing peptides. Sequence-dependent product profiles are obtained, and the abundance of deamidation-linked racemization is examined. Finally, comparisons are made between Gln deamidation which is relatively poorly understood, and asparagine deamidation which has been more thoroughly studied.
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DOI：
10.1073/pnas.78.3.1356
发表时间：
1981-01-01
期刊：
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES
影响因子：
--
作者：
LORAND, L;HSU, LKH;RAFFERTY, NS
通讯作者：
RAFFERTY, NS
影响因子：
4.4
作者：
Hains, Peter G.;Truscott, Roger J. W.
通讯作者：
Truscott, Roger J. W.
影响因子：
3.9
作者：
Lindner, H;Helliger, W
通讯作者：
Helliger, W
DOI：
10.1016/j.pbiomolbio.2014.02.004
发表时间：
2014-07
影响因子：
3.8
作者：
Lampi KJ;Wilmarth PA;Murray MR;David LL
通讯作者：
David LL
影响因子：
3.7
作者：
PATEL, K;BORCHARDT, RT
通讯作者：
BORCHARDT, RT