The analysis of alpha-1-antitrypsin glycosylation with direct LC-MS/MS.
The analysis of alpha-1-antitrypsin glycosylation with direct LC-MS/MS.
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使用直接 LC-MS/MS 分析 Alpha-1-抗胰蛋白酶糖基化
DOI：
10.1002/elps.201700426
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发表时间：
2018-09
期刊：
影响因子：
2.9
通讯作者：
Yao Z
中科院分区：
文献类型：
--
作者：
Yin H;An M;So PK;Wong MY;Lubman DM;Yao Z
建立了一种基于LC-MS/MS的方法来区分糖肽的核心岩藻糖基化和触角岩藻糖基化。糖基化位点(异质性)和每个位点的多个可能的糖链占有率(微异质性)都可以通过完整的糖肽分析来解决。血清糖蛋白α-1抗胰蛋白酶(A1AT)同时含有核岩藻糖基化和触角岩藻糖基化糖基。唾液酸酶用于去除唾液酸，以简化糖基化的微观异质性，并增强具有相似糖结构的糖肽的MS信号。β1-3，4半乳糖苷酶用于区分核心岩藻糖基化和触角岩藻糖基化。研究发现，源内解离严重影响低丰度糖链修饰糖肽的鉴定和定量。因此，质谱仪的设置被优化，以最大限度地减少源内解离。在pGlyco软件注释和人工检查的帮助下，糖肽的鉴定采用了三步质谱裂解策略。用于初始糖肽裂解的碰撞能量被发现对于改进对氧离子的检测和更好地选择Y1离子(肽+GlcNAc)是至关重要的。结构分析表明，A1AT糖肽的3个糖基化位点都含有复杂的N-糖链结构：Asn70位点含有不含岩藻糖基化的双触角多糖；Asn107位点含有同时具有核心和触角岩藻糖基化的双、三和四触角多糖；Asn271位点含有同时具有核心和触角岩藻糖基化的双触角和三触角多糖。Asn107的核心岩藻糖基化和触角岩藻糖化的相对强度与A1AT蛋白相似，说明Asn107的糖基化水平远高于其他两个位点。
A liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based methodology has been developed to differentiate core- and antennary-fucosylated glycosylation of glycopeptides. Both the glycosylation sites (heterogeneity) and multiple possible glycan occupancy at each site (microheterogeneity) can be resolved via intact glycopeptide analysis. The serum glycoprotein alpha-1-antitrypsin (A1AT) which contains both core- and antennary-fucosylated glycosites was used in this study. Sialidase was used to remove the sialic acids in order to simplify the glycosylation microheterogeneity and to enhance the MS signal of glycopeptides with similar glycan structures. β1-3,4 galactosidase was used to differentiate core- and antennary-fucosylation. In-source dissociation was found to severely affect the identification and quantification of glycopeptides with low abundance glycan modification. The settings of the mass spectrometer were therefore optimized to minimize the in-source dissociation. A three-step mass spectrometry fragmentation strategy was used for glycopeptide identification, facilitated by pGlyco software annotation and manual checking. The collision energy used for initial glycopeptide fragmentation was found to be crucial for improved detection of oxonium ions and better selection of Y1 ion (peptide+GlcNAc). Structural assignments revealed that all 3 glycosylation sites of A1AT glycopeptides contain complex N-glycan structures: site Asn70 contains biantennary glycans without fucosylation; site Asn107 contains bi-, tri- and tetra-antennary glycans with both core- and antennary-fucosylation; site Asn271 contains bi- and tri-antennary glycans with both core- and antennary-fucosylation. The relative intensity of core- and antennary-fucosylation on Asn107 was similar to that of the A1AT protein indicating that the glycosylation level of Asn107 is much larger than the other 2 sites.
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影响因子：
3.7
作者：
Comunale MA;Rodemich-Betesh L;Hafner J;Wang M;Norton P;Di Bisceglie AM;Block T;Mehta A
通讯作者：
Mehta A
DOI：
10.1016/j.jchromb.2016.02.006
发表时间：
2016-10-01
影响因子：
3
作者：
Szigeti, Marton;Bondar, Judit;Guttman, Andras
通讯作者：
Guttman, Andras
影响因子：
2.9
作者：
Guttman, A
通讯作者：
Guttman, A
通过固相提取N连接的聚糖和含糖材料的肽对蛋白质糖基化的全面分析。
影响因子：
46.9
作者：
Sun S;Shah P;Eshghi ST;Yang W;Trikannad N;Yang S;Chen L;Aiyetan P;Höti N;Zhang Z;Chan DW;Zhang H
通讯作者：
Zhang H
pGlyco：使用 HCD-和 CID-MS/MS 和 MS3 鉴定完整 N-糖肽的管道。
影响因子：
4.6
作者：
Zeng WF;Liu MQ;Zhang Y;Wu JQ;Fang P;Peng C;Nie A;Yan G;Cao W;Liu C;Chi H;Sun RX;Wong CC;He SM;Yang P
通讯作者：
Yang P