Rapid Quantification of Peptide Oxidation Isomers From Complex Mixtures.
Rapid Quantification of Peptide Oxidation Isomers From Complex Mixtures.
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DOI：
10.1021/acs.analchem.9b05268
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发表时间：
2020-03-03
影响因子：
7.4
通讯作者：
Sharp JS
中科院分区：
文献类型：
--
作者：
Khaje NA;Sharp JS
羟基自由基蛋白质足迹（HRPF）是一种基于定量蛋白质不同区域氧化量的强有力的蛋白质形貌变化探测技术。虽然在肽水平上定量HRPF氧化相对常见且简单，但由于氧化对MS/MS碎片的影响以及大量复杂且仅部分色谱分离的异构肽氧化产物，在残留物水平上定量具有挑战性。通过电子转移解离串联质谱法（ETD MS/MS）在残基水平上对异构肽氧化产物（其中肽序列相同，但异构氧化产物在不同位点形成）进行HRPF定量，已在模型肽和HRPF产物中得到证实，但该方法受到反相色谱法部分分离氧化异构体的阻碍。这需要定制的MS/MS方法在其洗脱窗口内对所有异构氧化产物进行同等采样，从而大大增加了方法开发时间，并减少了单次LC-MS/MS运行中定量的氧化产物。在这里，我们提出了一种两性离子亲水相互作用毛细管色谱（ZIC-HILIC）方法，理想地共混所有异构体的肽氧化产物，同时分离不同的肽。这使我们能够使用在单个肽氧化异构体峰上的任何点获得的ETD MS/MS光谱相对定量肽氧化异构体，大大简化了数据采集和数据分析。
Hydroxyl radical protein footprinting (HRPF) is a powerful technique for probing changes in protein topography, based on quantifying the amount of oxidation of different regions of a protein. While quantification of HRPF oxidation at the peptide level is relatively common and straightforward, quantification at the residue level is challenging due to the influence of oxidation on MS/MS fragmentation, and the large number of complex and only partially chromatographically resolved isomeric peptide oxidation products. HRPF quantification of isomeric peptide oxidation products (where the peptide sequence is the same but isomeric oxidation products are formed at different sites) at the residue level by electron transfer dissociation tandem mass spectrometry (ETD MS/MS) has been demonstrated in both model peptides and HRPF products, but the method is hampered by the partial separation of oxidation isomers by reversed phase chromatography. This requires custom MS/MS methods to equally sample all isomeric oxidation products across their elution window, greatly increasing method development time and reducing the oxidation products quantified in a single LC-MS/MS run. Here we present a zwitterionic hydrophilic interaction capillary chromatography (ZIC-HILIC) method to ideally co-elute all isomeric peptide oxidation products while separating different peptides. This allows us to relatively quantify peptide oxidation isomers using an ETD MS/MS spectrum acquired at any point across the single peptide oxidation isomer peak, greatly simplifying data acquisition and data analysis.
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DOI：
10.1016/s0021-9673(00)96972-3
发表时间：
1990-01-19
期刊：
JOURNAL OF CHROMATOGRAPHY
影响因子：
--
作者：
ALPERT, AJ
通讯作者：
ALPERT, AJ
DOI：
10.1016/j.ymeth.2015.02.018
发表时间：
2015-11-01
期刊：
Methods (San Diego, Calif.)
影响因子：
--
作者：
Calabrese AN;Ault JR;Radford SE;Ashcroft AE
通讯作者：
Ashcroft AE
DOI：
10.1007/s13361-013-0719-5
发表时间：
2013-11
影响因子：
3.2
作者：
Li, Xiaoyan;Li, Zixuan;Xie, Boer;Sharp, Joshua S.
通讯作者：
Sharp, Joshua S.
影响因子：
7.4
作者：
Vahidi, Siavash;Stocks, Bradley B.;Konermann, Lars
通讯作者：
Konermann, Lars
影响因子：
2.9
作者：
Li X;Grant OC;Ito K;Wallace A;Wang S;Zhao P;Wells L;Lu S;Woods RJ;Sharp JS
通讯作者：
Sharp JS