Center for Intelligent Chemical Instrumentation, Department of Chemistry and Biochemistry, Ohio University, Athens, OH 45701, USA.
J Am Soc Mass Spectrom. 2011 Sep;22(9):1610-21. doi: 10.1007/s13361-011-0170-4. Epub 2011 Jun 9.
Our previous study showed that selenamide reagents such as ebselen and N-(phenylseleno)phthalimide (NPSP) can be used for selective and rapid derivatization of protein/peptide thiols in high conversion yield. This paper reports the systematic investigation of MS/MS dissociation behaviors of selenamide-derivatized peptide ions upon collision induced dissociation (CID) and electron transfer dissociation (ETD). In the positive ion mode, derivatized peptide ions exhibit tag-dependent CID dissociation pathways. For instance, ebselen-derivatized peptide ions preferentially undergo Se-S bond cleavage upon CID to produce a characteristic fragment ion, the protonated ebselen (m/z 276), which allows selective identification of thiol peptides from protein digest as well as selective detection of thiol proteins from protein mixture using precursor ion scan (PIS). In contrast, NPSP-derivatized peptide ions retain their phenylselenenyl tags during CID, which is useful in sequencing peptides and locating cysteine residues. In the negative ion CID mode, both types of tags are preferentially lost via the Se-S cleavage, analogous to the S-S bond cleavage during CID of disulfide-containing peptide anions. In consideration of the convenience in preparing selenamide-derivatized peptides and the similarity of Se-S of the tag to the S-S bond, we also examined ETD of the derivatized peptide ions to probe the mechanism for electron-based ion dissociation. Interestingly, facile cleavage of Se-S bond occurs to the peptide ions carrying either protons or alkali metal ions, while backbone cleavage to form c/z ions is severely inhibited. These results are in agreement with the Utah-Washington mechanism proposed for depicting electron-based ion dissociation processes.
我们之前的研究表明，硒酰胺试剂，如依布硒啉和 N-（苯硒基）邻苯二甲酰亚胺（NPSP），可用于选择性和快速衍生高转化率的蛋白质/肽巯基。本文报道了对硒酰胺衍生肽离子在碰撞诱导解离（CID）和电子转移解离（ETD）下的 MS/MS 解离行为的系统研究。在正离子模式下，衍生化的肽离子表现出依赖于标签的 CID 解离途径。例如，依布硒啉衍生肽离子在 CID 时优先发生 Se-S 键断裂，产生特征碎片离子，即质子化的依布硒啉（m/z 276），这允许从蛋白质消化物中选择性鉴定巯基肽，以及使用前体离子扫描（PIS）从蛋白质混合物中选择性检测巯基蛋白。相比之下，NPSP 衍生肽离子在 CID 过程中保留其苯硒基标签，这在测序肽和定位半胱氨酸残基方面很有用。在负离子 CID 模式下，两种类型的标签都通过 Se-S 断裂优先丢失，类似于含二硫键的肽阴离子 CID 过程中的 S-S 键断裂。考虑到制备硒酰胺衍生肽的便利性和标签的 Se-S 与 S-S 键的相似性，我们还研究了衍生化肽离子的 ETD，以探究基于电子的离子解离机制。有趣的是，携带质子或碱金属离子的肽离子容易发生 Se-S 键断裂，而形成 c/z 离子的骨架断裂则受到严重抑制。这些结果与用于描述基于电子的离子解离过程的犹他-华盛顿机制一致。
