Chemical derivatization of peptide carboxyl groups for highly efficient electron transfer dissociation.
Chemical derivatization of peptide carboxyl groups for highly efficient electron transfer dissociation.
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DOI：
10.1007/s13361-013-0701-2
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发表时间：
2013-11
影响因子：
3.2
通讯作者：
Smith, Lloyd M.
中科院分区：
文献类型：
--
作者：
Frey, Brian L.;Ladror, Daniel T.;Sondalle, Samuel B.;Krusemark, Casey J.;Jue, April L.;Coon, Joshua J.;Smith, Lloyd M.
关键词：
胰蛋白酶肽的羧基用叔胺或季胺标记试剂衍生化，以产生更高电荷的肽离子，其通过电子转移解离（ETD）有效地片段化。所有肽羧基天冬氨酸和谷氨酸侧链以及C-末端衍生化的平均反应效率为99%。这种几乎完全的标记避免了由于部分标记的产物而使复杂的肽混合物变得更加复杂，并且它允许在数据库搜索期间使用静态修饰。烷基叔胺被认为是最佳的标记试剂中的四种类型的测试。衍生化肽的电荷状态明显更高：牛血清白蛋白（BSA）的修饰胰蛋白酶消化物产生约90%的z > 2的前体离子，而未修饰样品的前体离子不到40%。修饰的肽离子的电荷密度增加产生高效的ETD片段化，导致许多额外的肽鉴定和更高的序列覆盖率（例如修饰的70%，而未修饰的BSA仅为43%）。这种标记策略的效用被证明对从酵母细胞分离的核糖体蛋白的胰蛋白酶消化。该样品的肽衍生化产生了鉴定的蛋白质数量的增加，这些蛋白质的序列覆盖度增加>50%，并且肽光谱匹配的数量加倍。这种羧基衍生化策略大大提高了从胰蛋白酶消化酶的ETD-MS/MS获得的蛋白质组覆盖率，我们预计它也将增强翻译后修饰的识别和定位。
The carboxyl groups of tryptic peptides were derivatized with a tertiary or quaternary amine labeling reagent to generate more highly charged peptide ions that fragment efficiently by electron transfer dissociation (ETD). All peptide carboxyl groups—aspartic and glutamic acid side-chains as well as C-termini—were derivatized with an average reaction efficiency of 99%. This nearly complete labeling avoids making complex peptide mixtures even more complex due to partially-labeled products, and it allows the use of static modifications during database searching. Alkyl tertiary amines were found to be the optimal labeling reagent among the four types tested. Charge states are substantially higher for derivatized peptides: a modified tryptic digest of bovine serum albumin (BSA) generates ∼90% of its precursor ions with z > 2, compared to less than 40% for the unmodified sample. The increased charge density of modified peptide ions yields highly efficient ETD fragmentation, leading to many additional peptide identifications and higher sequence coverage (e.g. 70% for modified versus only 43% for unmodified BSA). The utility of this labeling strategy was demonstrated on a tryptic digest of ribosomal proteins isolated from yeast cells. Peptide derivatization of this sample produced an increase in the number of identified proteins, a >50% increase in the sequence coverage of these proteins, and a doubling of the number of peptide spectral matches. This carboxyl derivatization strategy greatly improves proteome coverage obtained from ETD-MS/MS of tryptic digests, and we anticipate that it will also enhance identification and localization of post-translational modifications.
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影响因子：
7.4
作者：
Kjeldsen, Frank;Giessing, Anders M. B.;Jensen, Ole N.
通讯作者：
Jensen, Ole N.
DOI：
10.1016/j.jasms.2006.10.005
发表时间：
2007-03-01
影响因子：
3.2
作者：
Jones, Jace W.;Sasaki, Tomikazu;Turecek, Frantisek
通讯作者：
Turecek, Frantisek
影响因子：
1.8
作者：
Frey BL;Krusemark CJ;Ledvina AR;Coon JJ;Belshaw PJ;Smith LM
通讯作者：
Smith LM
影响因子：
4.4
作者：
Hsu, JL;Huang, SY;Chen, SH
通讯作者：
Chen, SH
影响因子：
7.4
作者：
Krokhin, Oleg V.;Spicer, Vic
通讯作者：
Spicer, Vic