Improved tandem mass spectrometric characterization of 3-nitrotyrosine sites in peptides
Improved tandem mass spectrometric characterization of 3-nitrotyrosine sites in peptides
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DOI：
10.1002/rcm.2676
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发表时间：
2006-01-01
影响因子：
2
通讯作者：
Gevaert, Kris
中科院分区：
文献类型：
--
作者：
Ghesquiere, Bart;Goethals, Marc;Gevaert, Kris
硝化酪氨酸很容易通过还原步骤转化为它们的氨基酪氨酸等价物。我们在这里表明，这种转化可以很容易地在非硝化肽的背景中辨别3-氨基酪氨酸肽。此外，氨基酪氨酸肽在单质谱（MS）模式下更稳定，使得肽质量图更容易解释。3-硝基酪氨酸和3-氨基酪氨酸肽的一个重要警告是它们在碰撞诱导解离（CID）时缺乏有效的碎片化，在后一种肽的情况下，也会产生意想不到的，偏离含有氨基酪氨酸残基的碎片离子的同位素模式。最终的结果是序列数据库搜索变得令人生畏，因为正确的肽经常被遗漏，因为使用了不充分和/或不准确的肽片段。我们证明了一个简单的乙酰化步骤，阻断所有胺（包括氨基酪氨酸），产生的肽经过CID广泛的骨干断裂，因此很容易在数据库中识别。与使用相同量的物质直接分析硝化肽相比，我们的程序通过在四硝基甲烷硝化的牛血清白蛋白（BSA）中绘制的硝化事件数量增加一倍来进一步说明。总之，我们在这里说明了这两步过程，血红素介导的还原和乙酰化，可以用于更有效地表征蛋白质结合的硝化酪氨酸。版权所有(c) 2006约翰威利父子有限公司
Nitrated tyrosines are easily converted into their aminotyrosine equivalents by a reduction step. We here show that this conversion can be exploited to readily discern 3-aminotyrosine peptides in a background of non-nitrated peptides. Furthermore, aminotyrosine peptides are more stable in single mass spectrometry (MS) mode rendering peptide mass maps easier to interpret. One significant caveat of both 3-nitrotyrosine and 3-aminotyrosine peptides is their lack of efficient fragmentation upon collision-induced dissociation (CID) which, in the case of the latter peptides, also produces unexpected, deviating isotopic patterns of fragment ions containing the aminotyrosine residue. The net result is that sequence database searching becomes daunting as the correct peptide is frequently missed since insufficient and/or inaccurate peptide fragments are used. We show that a simple acetylation step, blocking all amines (including aminotyrosine), produces peptides that undergo extensive backbone fragmentation by CID and are thus easily identifiable in databases. Our procedure is additionally illustrated by doubling the number of nitration events mapped in tetranitromethane-nitrated bovine serum albumin (BSA) as compared to a direct analysis of the nitrated peptides using the same amount of material. In conclusion, we here illustrate that this twostep process, heme-mediated reduction and acetylation, can be used for more efficient characterization of protein-bound nitrated tyrosines. Copyright (c) 2006 John Wiley & Sons, Ltd.