Chemistry Department, M.V. Lomonosov Moscow State University , Leninskie Gory 1/3, Moscow, 119991, Russia.
Anal Chem. 2014 Jul 15;86(14):7017-22. doi: 10.1021/ac501200h. Epub 2014 Jun 30.
An efficient approach to easy and reliable differentiation between isomeric leucine and isoleucine in peptide sequencing utilizes multistage electron transfer dissociation and higher energy collision activated dissociation in the Orbitrap Fusion mass spectrometer. The MS(3) method involves production and isolation of primary odd-electron z(•) ions, followed by radical site initiation of their fragmentation with formation of w-ions, characteristic of the isomeric amino acid residues. Six natural nontryptic peptides isolated from the secretion of frog Rana ridibunda were studied. Their lengths were in the range between 15 and 37 amino acids and the number of targeted isomeric (Leu/Ile) residues varied between 1 and 7. The experiments were successful in all 22 cases of Leu/Ile residues, leaving no doubts in identification. The method is extremely selective as the targeted w-ions appear to be the most intense in the spectra. The proposed approach may be incorporated into shotgun proteomics algorithms and allows for the development of an exclusively mass spectrometric method for automated complete de novo sequencing of various peptides and proteins.
一种高效的方法，可轻松可靠地区分肽测序中异构的亮氨酸和异亮氨酸，该方法利用多级电子转移解离和轨道阱 Fusion 质谱仪中的更高能量碰撞激活解离。MS(3) 方法涉及初级奇数电子 z(•)离子的生成和分离，随后进行自由基引发碎裂，形成 w-离子，这是异构氨基酸残基的特征。从蛙 Rana ridibunda 的分泌物中分离出的六种天然非胰蛋白酶肽进行了研究。它们的长度在 15 到 37 个氨基酸之间，目标异构（亮氨酸/异亮氨酸）残基的数量在 1 到 7 之间不等。在所有 22 个亮氨酸/异亮氨酸残基的情况下，实验均获得成功，鉴定结果毫无疑问。该方法具有极高的选择性，因为靶向的 w-离子在光谱中似乎是最强的。该方法可纳入鸟枪法蛋白质组学算法，并允许开发一种完全依赖质谱的方法，用于自动完成各种肽和蛋白质的从头测序。
