signature=d49b5fbe5c21e280a1d3f44ac9bb9cb6,Tandem mass spectra of glycan substructures enable the mu...-优快云博客

研究中，通过创建非还原末端糖基化结构的串联质谱碎片库，提出了一种识别和确认这些糖基化子结构的方法。使用离子阱质谱分析分离的寡糖，并通过MS2和MS3碎片化确定糖基化子结构，尤其在负离子模式下，对于区分同分异构糖结构非常有用。该策略已应用于复杂N-和O-糖链中硫酸化、血型抗原和唾液酸连接方式的鉴定。

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摘要：

RATIONALEGlycosylation of proteins and lipids affects many biological processes, such as host–pathogen interactions, cell communication, and initiation of the immune responses. Terminal glycan substructures, or determinants, often govern the function or recognition of the carrier glycoconjugate and modulate these processes. In this study we describe a strategy using multistage mass spectrometry to identify and confirm these glycan substructures.METHODSAn online tandem mass spectrometry (MS2) spectral fragment library of glycan substructures that typically occur at the non-reducing terminus of glycoconjugates was created to enable the easier identification and confirmation of glycan determinants on oligosaccharides released from glycoproteins. Oligosaccharides were separated by porous graphitized carbon capillary chromatography and analysed by ion trap MS. Candidate product ions that constitute the glycan substructure mass were identified in the MS2 product ion spectrum, and used as the precursor ion for subsequent MS3 fragmentation. The resulting MS3 spectrum was matched against the MS2 spectral fragment library to identify the glycan substructure(s) that comprise the parent oligosaccharide.RESULTSThirty biologically important terminal glycan determinants commonly observed on glycoconjugates were fragmented by positive and negative ion mass spectrometry and the MS2 product ion masses manually annotated and stored in the UniCarb-DB online database. Negative ion tandem mass spectra were especially useful in assigning isobaric glycan structures. We have applied this strategy for the identification of the sulphation, blood group antigens and sialic acid linkages on complex N-and O-glycans released from glycoproteins.CONCLUSIONSWe show the potential of these glycan substructure MS2 spectra in the negative ionization mode to facilitate the assignment of determinants on N- and O-linked glycans released from glycoproteins. Comparing the structural feature ions of known glycan reference substructures assists in the annotation of complex glycan product ion spectra, and can remove the need for other orthogonal confirmation analyses such as sequential glycosidase digestion. Copyright 2013 John Wiley & Sons, Ltd.

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