Gardner Myles W, Brodbelt Jennifer S
Department of Chemistry and Biochemistry, The University of Texas at Austin, Austin, Texas 78712, USA.
J Am Soc Mass Spectrom. 2008 Mar;19(3):344-57. doi: 10.1016/j.jasms.2007.11.006. Epub 2007 Nov 19.
The dissociation of intermolecularly crosslinked peptides was evaluated for a series of peptides with proline or aspartic acid residues positioned adjacent to the crosslinking sites (lysine residues). The peptides were crosslinked with either disuccinimidyl suberate (DSS) or disuccinimidyl L-tartrate (DST), and the influence of proline and aspartic acid residues on the fragmentation patterns were investigated for precursor ions with and without a mobile proton. Collisionally activated dissociation (CAD) spectra of aspartic acid-containing crosslinked peptide ions, doubly-charged with both protons sequestered, were dominated by cleavage C-terminal to the Asp residue, similar to that of unmodified peptides. The proline-containing crosslinked peptides exhibited a high degree of internal ion formation, with the resulting product ions having an N-terminal proline residue. Upon dissociation of the doubly-charged crosslinked peptides, twenty to fifty percent of the fragment ion abundance was accounted for by multiple cleavage products. Crosslinked peptides possessing a mobile proton yielded almost a full series of b- and y-type fragment ions, with only proline-directed fragments still observed at high abundances. Interestingly, the crosslinked peptides exhibited a tendency to dissociate at the amide bond C-terminal to the crosslinked lysine residue, relative to the N-terminal side. One could envision updating computer algorithms to include these crosslinker specific product ions--particularly for precursor ions with localized protons--that provide complementary and confirmatory information, to offer more confident identification of both the crosslinked peptides and the location of the crosslink, as well as affording predictive guidelines for interpretation of the product-ion spectra of crosslinked peptides.
对于一系列在交联位点（赖氨酸残基）相邻位置含有脯氨酸或天冬氨酸残基的肽段，评估了分子间交联肽段的解离情况。这些肽段用辛二酸二琥珀酰亚胺酯（DSS）或L-酒石酸二琥珀酰亚胺酯（DST）进行交联，并研究了脯氨酸和天冬氨酸残基对带有和不带有移动质子的前体离子碎裂模式的影响。对于两个质子都被螯合的双电荷含天冬氨酸交联肽离子的碰撞诱导解离（CAD）光谱，以天冬氨酸残基C端的裂解为主，类似于未修饰肽段。含脯氨酸的交联肽段表现出高度的内部离子形成，产生的产物离子带有N端脯氨酸残基。在双电荷交联肽段解离时，20%至50%的碎片离子丰度由多个裂解产物构成。具有移动质子的交联肽段几乎产生了完整系列的b型和y型碎片离子，只有脯氨酸导向的碎片在高丰度下仍被观察到。有趣的是，相对于N端，交联肽段倾向于在交联赖氨酸残基C端的酰胺键处解离。可以设想更新计算机算法，将这些交联剂特异性产物离子纳入其中——特别是对于带有局部质子的前体离子——这些离子可提供互补和确认信息，以便更可靠地鉴定交联肽段及其交联位置，同时为交联肽段的产物离子光谱解释提供预测指导。
