Mass spectrometric glycan rearrangements

Mass spectrometric rearrangement reactions have been reported for a large variety of compounds such as peptides, lipids, and carbohydrates. In the case of carbohydrates this phenomenon has been described as internal residue loss. Resulting fragment ions may be misinterpreted as fragments arising from conventional glycosidic bond cleavages, which may result in incorrect structural assignment. Therefore, awareness of the occurrence of glycan rearrangements is important for avoiding misinterpretation of tandem mass spectra. In this review mass spectrometric rearrangements of both derivatized and underivatized (native) oligosaccharide structures are discussed. Similar phenomena have been reported for glycopeptides, labeled glycan structures and other biomolecules containing a carbohydrate part. Rearrangements in oligosaccharides and glycoconjugates have been observed with different types of mass spectrometers. Most of the observed carbohydrate rearrangement reactions appear to be linked to the presence of a proton. Hence, tandem mass spectrometric analysis of alkali adducts or deprotonated ions often prevents rearrangement reactions, while they may happen with high efficacy with protonated glycoconjugates.     

DerivatizationStrategyforGlycomics

Glycomics is the systematic identification and characterization of diverse glycans for their structure, function, activity, and quantity. Global profiling of glycomes of cancer cells, body fluids, or tissues has been considered as an innovative and comprehensive approach to discover and identify biomarkers for various diseases. Neutralization of carboxylic acid is an important means to avoid sialic acid dissociation when sialylated oligosaccharides were analyzed by matrix-assisted desorption/ionization mass spectrometry (MALDI-MS). In this presentation, I will first discuss a simple and rapid method to modify sialic acids at sialylated oligosaccharides in presence of methylamine and (7-azabenzotriazol-1-yloxy) trispyrroli- dinophosphonium hexafluorophosphate (PyAOP). After methylamidation, sialylated oligosaccharides can be analyzed by MALDI-MS without loss of sialic acid moiety. The ESI-MS and MALDI-MS analysis of both 3’- and 6’-sialyllactose derivatives indicated that the quantitatively conversion of sialic acids was achieved, regardless of their linkage types under the optimal derivatization conditions. This derivatization strategy was further validated with the N-glycans released from three standard glycoproteins (Fetuin, human acid glycoprotein and bovine acid glycoprotein) containing different types of complex oligosaccharides. Using this derivatization method, the N-glycans of sera from different species (human, mouse and rat) were successfully characterized by MALDI-MS. Due to the mild reaction conditions, the modification in sialic acid residues can be retained. This improvement makes it possible to detect the sialylated oligosaccharides containing O-acetylated sialic acid moieties using MALDI-MS in positive ion mode. In the meantime, the development of glycomics is limited by the ability to identify and quantify large numbers of glycans in complex biological samples, which is only partially achieved by current glycomics approaches. I will also present a strategy termed targeted glycomics enabling highly sensitive and consistent identification and quantification of target sets of glycans across multiple samples. The strategy was first applied to the analysis of glycans released from ribonuclease B. A linear dynamic range of three orders of magnitude (0.1-100 ng ribonuclease B) was achieved for the detection of all five glycans. The method exhibited excellent sensitivity, evident by the detection of limit of 100 attomolar for Man9GlcNAc2. The strategy was further demonstrated by characterizing the glycome of pancreas cancer cells. More than 60 N-glycans were identified and detected for about five million cells.     

Real-time,label-free characterization of oligosaccharide-binding proteins using carbohydrate microarrays and an ellipsometry-based biosensor

Carbohydrates present on cell surfaces mediate cell behavior through interactions with other biomolecules. Due to their structural complexity, diversity, and heterogeneity, it is difficult to fully characterize a variety of carbohydrates and their binding partners. As a result, novel technologies for glycomics applications have been developed, including carbohydrate microarrays and label-free detection methods. In this paper, we report using the combination of oligosaccharide microarrays and the label-free oblique-incidence reflectivity difference microscopy for real-time characterization of oligosaccharide-binding proteins. Aminated human milk oligosaccharides were immobilized on epoxy-coated glass substrates as microarrays for reactions with Family 1 of solute-binding proteins from Bifidobacterium longum subsp. infantis (B. infantis). Binding affinities of these protein–oligosaccharide interactions showed preferences of Family 1 of solute-binding proteins to host glycans, which helps in characterizing the complex process of human milk oligosaccharides foraging by B. infantis.     

Glycomics and glycoproteomics of viruses: Mass spectrometry applications and insights toward structure–function relationships

The advancement of viral glycomics has paralleled that of the mass spectrometry glycomics toolbox. In some regard the glycoproteins studied have provided the impetus for this advancement. Viral proteins are often highly glycosylated, especially those targeted by the host immune system. Glycosylation tends to be dynamic over time as viruses propagate in host populations leading to increased number of and/or “movement” of glycosylation sites in response to the immune system and other pressures. This relationship can lead to highly glycosylated, difficult to analyze glycoproteins that challenge the capabilities of modern mass spectrometry. In this review, we briefly discuss five general areas where glycosylation is important in the viral niche and how mass spectrometry has been used to reveal key information regarding structure–function relationships between viral glycoproteins and host cells. We describe the recent past and current glycomics toolbox used in these analyses and give examples of how the requirement to analyze these complex glycoproteins has provided the incentive for some advances seen in glycomics mass spectrometry. A general overview of viral glycomics, special cases, mass spectrometry methods and work-flows, informatics and complementary chemical techniques currently used are discussed. © 2020 The Authors. Mass Spectrometry Reviews published by John Wiley & Sons Ltd. Mass Spec Rev     

Chip‐mass spectrometry for glycomic studies

The introduction of micro‐ and nanochip front end technologies for electrospray mass spectrometry addressed a major challenge in carbohydrate analysis: high sensitivity structural determination and heterogeneity assessment in high dynamic range mixtures of biological origin. Chip‐enhanced electrospray ionization was demonstrated to provide reproducible performance irrespective of the type of carbohydrate, while the amenability of chip systems for coupling with different mass spectrometers greatly advance the chip/MS technique as a versatile key tool in glycomic studies. A more accurate representation of the glycan repertoire to include novel biologically‐relevant information was achieved in different biological sources, asserting this technique as a valuable tool in glycan biomarker discovery and monitoring. Additionally, the integration of various analytical functions onto chip devices and direct hyphenation to MS proved its potential for glycan analysis during the recent years, whereby a new analytical tool is on the verge of maturation: lab‐on‐chip MS glycomics. The achievements until early beginning of 2007 on the implementation of chip‐ and functional integrated chip/MS in systems glycobiology studies are reviewed here. © 2009 Wiley Periodicals, Inc., Mass Spec Rev 28:223–253, 2009     

On-line separations combined with MS for analysis of glycosaminoglycans

The glycosaminoglycan (GAG) family of polysaccharides includes the unsulfated hyaluronan and the sulfated heparin, heparan sulfate, keratan sulfate, and chondroitin/dermatan sulfate. GAGs are biosynthesized by a series of enzymes, the activities of which are controlled by complex factors. Animal cells alter their responses to different growth conditions by changing the structures of GAGs expressed on their cell surfaces and in extracellular matrices. Because this variation is a means whereby the functions of the limited number of protein gene products in animal genomes is elaborated, the phenotypic and functional assessment of GAG structures expressed spatially and temporally is an important goal in glycomics. On-line mass spectrometric separations are essential for successful determination of expression patterns for the GAG compound classes due to their inherent complexity and heterogeneity. Options include size exclusion, anion exchange, reversed phase, reversed phase ion pairing, hydrophilic interaction, and graphitized carbon chromatographic modes and capillary electrophoresis. This review summarizes the application of these approaches to on-line MS analysis of the GAG classes.     

Structural investigation of bacterial lipopolysaccharides by mass spectrometry and tandem mass spectrometry

Mass spectrometric studies are now playing a leading role in the elucidation of lipopolysaccharide (LPS) structures through the characterization of antigenic polysaccharides, core oligosaccharides and lipid A components including LPS genetic modifications. The conventional MS and MS/MS analyses together with CID fragmentation provide additional structural information complementary to the previous analytical experiments, and thus contribute to an integrated strategy for the simultaneous characterization and correct sequencing of the carbohydrate moiety. © 2010 Wiley Periodicals, Inc., Mass Spec Rev 29:606–650, 2010     

Structural glycomics using hydrophilic interaction chromatography (HILIC) with mass spectrometry

Hydrophilic interaction chromatography (HILIC) with mass spectrometry is a versatile technique for structural glycomics. Glycans are retained by hydrogen bonding, ionic interactions, and dipole-dipole interactions. Glycopeptides as well as glycans with various modifications and reducing-end labels can be efficiently separated, which often results in the resolution of isobaric species. Chromatography is usually performed with solvent mixtures of organic modifier (often acetonitrile) and volatile (acidic) buffer which are suitable for online-electrospray ionization-mass spectrometry. When performed at the nano-scale, this results in a detection limit for oligosaccharides of approximately 1 femtomol. Alternatively, glycans may be analyzed by offline-MALDI-MS(/MS) in both negative-ion mode and positive-ion mode, which allows the registration of informative fragment ion spectra from deprotonated species and sodium adducts, respectively. (c) 2009 Wiley Periodicals, Inc., Mass Spec Rev 28:192-206, 2009.     

Chemical cross-linking and mass spectrometry to map three-dimensional protein structures and protein-protein interactions

Closely related to studying the function of a protein is the analysis of its three-dimensional structure and the identification of interaction sites with its binding partners. An alternative approach to the high-resolution methods for three-dimensional protein structure analysis, such as X-ray crystallography and NMR spectroscopy, consists of covalently connecting two functional groups of the protein(s) under investigation. The location of the created cross-links imposes a distance constraint on the location of the respective side chains and allows one to draw conclusions on the three-dimensional structure of the protein or a protein complex. Recently, chemical cross-linking of proteins has been combined with a mass spectrometric analysis of the created cross-linked products. This review article describes the most popular cross-linking reagents for protein structure analysis and gives an overview of the different available strategies that employ chemical cross-linking and different mass spectrometric techniques. The challenges for mass spectrometry caused by the enormous complexity of the cross-linking reaction mixtures are emphasized. The various approaches described in the literature to facilitate the mass spectrometric detection of cross-linked products as well as computer software for data analyses are reviewed.     

美国应用生物系统公司串联液质系统--食品安全的卫士

高效液相色谱-串联质谱联用技术在近几年发展迅猛，已成为在复杂体系中进行微量组分的筛选、定性和定量极其可靠、快速和准确的分析手段。受到了学术界和分析测试行业的普遍青睐，并在生命科学、药学、医学、环境与健康等领域得到广泛应用。本文论述了美国应用生物系统公司串联液质系统用于食品安全领域关键的技术要点，比如三级四极杆质谱技术擅长快速定量分析，就好比是“长了眼睛的分析检测技术”，不需要液相色谱的完全分离，提高了方法建立与分析检测的速度，并拓展了分析测试的检测下限；同时。线性离子阱技术可提供高质量的化合物碎片离子全扫描图谱以及多级串联质谱功能，大大增强了系统的定性功能。而两种技术的完美组合即“串联四极杆-线性离子阱融合质谱技术”不仅出色地完成高灵敏度、高选择性的定量分析，同时可以获得增强性化和物结构碎片图谱，使定性、定量工作一次完成，大大提高了食品安全中分析检测的通量．本文介绍了这种新技术的基本原理及其在食品安全中的经典应用。     

Characterization of proteins by ambient mass spectrometry

Proteins play important roles in living systems and are topics of many fundamental and applied research projects. With the introduction of electrospray ionization and matrix‐assisted laser desorption/ionization for analysis of biomacromolecules in the late 1980s, mass spectrometry has become an important tool for characterization of proteins. Characterization of proteins in raw samples by these mass spectrometric techniques, however, usually requires extensive sample pretreatment. Ambient ionization techniques are new mass spectrometric techniques that allow direct analysis of samples with no or little sample preparation. Can these techniques facilitate or even eliminate sample preparation for mass spectrometric analysis of proteins? Apart from sample preparation, do these techniques offer any new features for characterization of proteins as compared with conventional ESI or MALDI? Recent advances in characterization of proteins by ambient mass spectrometry are summarized and commented in this article. © 2011 Wiley Periodicals, Inc. Mass Spec Rev 31:437–447, 2012     

生物质谱在基因组学研究中的应用

综述生物质谱用于分析核糖核酸的应用进展。概述电喷雾电离质谱(ESI-MS)和基质辅助激光解吸电离质谱(MALDI-MS)的原理及它们在核酸分析中的应用。总结质谱用于单核苷酸多态性分型分析(SNPgenotyping);对短的串联重复序列(STR)的分析;对寡核苷酸片断的序列分析等三个方面的研究成果。提出质谱用于基因组学研究存在的问题,并展望生物质谱未来的发展方向。     

有机化合物表面电离质谱技术

文章简述有机化合物表面电离质谱技术的发展和特性,介绍了三种电离类型:分子表面电离(MSI)、分解表面电离(DSI)和缔合表面电离(ASI),以及相对灵敏度与电子轰击电离灵敏度比较,离子流强度与样品分压、样品吸附时间、热发射表面温度的关系。讨论了含氮直链和环状有机化合物、碳氢化合物、含氧有机化合物和金属有机化合物表面电离质谱,列出了其中45种有机化合物的分子组成、电离能或出现电离能、强峰离子、相对灵敏度。     

Crosslinking combined with mass spectrometry for structural proteomics

The method of crosslinking combined with mass spectrometry is being gradually accepted as a technology enabling detailed structural information on proteins and protein complexes. Intrinsic challenges of the method, which have prevented its widespread use, are being progressively addressed by improvements in mass spectrometry instrumentation capabilities, by the development of new crosslinking reagents, and by the development of specialized software tools for processing of mass spectrometric crosslinking data. This review focuses on recent literature concerning the development of specialized crosslinking reagents and approaches for mass spectrometry‐based applications. Critical features of crosslinking reagents for optimum mass spectrometric performance, such as isotopic coding, cleavability, affinity groups, structure of the linkers, and reactive groups, are assessed. Requirements for the design of crosslinking reagents to make them well suited for mass spectrometric detection and analysis are summarized. © 2010 Wiley Periodicals, Inc. Mass Spec Rev 29:862–876, 2010     

Infrared multiphoton dissociation in quadrupole ion traps

The development of new ion activation techniques continues to be a dynamic area of scientific discovery, in part to complement the tremendous innovations in ionization methods that have allowed the mass spectrometric analysis of an enormous array of molecules. Ion activation/dissociation provides key information about ion structures, binding energies, and differentiation of isomers, as well as affording a primary means of identifying compounds in mixtures. Numerous new activation methods have emerged over the past two decades in an effort to develop alternatives to collisional activated dissociation, the gold standard for providing structurally diagnostic fragmentation patterns. Collisional activated dissociation does not always offer sufficiently high or controllable energy deposition, thus rendering it less useful for certain classes of molecules, such as large proteins or macromolecular complexes. Photodissociation is one of the most promising alternatives and is readily implemented in ion trapping and time‐of‐flight mass spectrometers. Photodissociation generally entails using a laser to irradiate ions with UV, visible, or IR photons, thus resulting in internal energy deposition based on the number and wavelengths of the photons. The activation process can be extremely rapid and efficient, as well as having the potential for high total energy deposition. This review describes infrared multiphoton dissociation in quadrupole ion trap mass spectrometry. A comparison of photodissociation and collisional activated dissociation is covered, in addition to some of the methods to increase photodissociation efficiency. Numerous applications of IRMPD are discussed as well, including ones related to the analysis of drugs, peptides, nucleic acids, and oligosaccharides. © 2009 Wiley Periodicals, Inc., Mass Spec Rev 28:390–424, 2009     

多肽及蛋白质质谱分析新进展

多肽和蛋白质的结构分析常用化学及生物化学降解法，近年来运用质谱法日益增多，本文简要综述质谱新技术，如快原子轰击电离、串联质谱、电喷雾质等应用于多肽和蛋白质分子量测定及序列分析的新进展。     

气相色谱-质谱法测定X胶囊的成分(英文)

建立了采用气相色谱-质谱法(GC/MS)确定X胶囊中内容物组成的方法。X胶囊中的内容物研磨成细粉后用乙醚提取,挥干乙醚后,溶于四氢呋喃进样。测定结果表明:样品溶液获得的GC/MS谱图数据与甾体化合物甲基睾丸素(甲睾)非常相似。通过对照发现样品的GC/MS谱图与甲睾标准品的GC/MS谱图完全相同。由此可以鉴定X胶囊样品为甲睾。     

Mass spectrometry analysis of the influenza virus

The role of mass spectrometry to probe characteristics of the influenza virus, and vaccine and antiviral drugs that target the virus, are reviewed. Genetic and proteomic approaches have been applied which incorporate high resolution mass spectrometry and mass mapping to genotype the virus and establish its evolution in terms of the primary structure of the surface protein antigens. A mass spectrometric immunoassay has been developed and applied to assess the structure and antigenicity of the virus in terms of the hemagglutinin antigen. The quantitation of the hemagglutinin antigen in vaccine preparations has also been conducted that is of importance to their efficacy. Finally, the characterization and quantitation of antiviral drugs against the virus, and their metabolites, have been monitored in blood, serum, and urine. The combined approaches demonstrate the strengths of modern mass spectrometric methods for the characterization of this killer virus. [This article was published online 10 September 2008. An error was subsequently identified. This notice is included in the online and print versions to indicate that both have been corrected 7 November 2008.]     

漂移时间离子淌度-四极杆-飞行时间串联质谱法分析寡糖同分异构体

分析糖类分子的结构是研究生物功能的必要前提，然而一种糖类常存在多种同分异构体，传统方法难以实现快速有效鉴别。本研究使用漂移时间离子淌度-四极杆-飞行时间串联质谱法将乙腈-水-甲酸溶液中的寡糖同分异构体分子经过电喷雾电离源电离，在漂移管内实现基于离子分子结构和带电荷数的分离，离子在四极杆中碎裂，最终被飞行时间质谱检测。寡糖同分异构体离子到达检测器的时间相差0.15~0.66 ms，能实现部分分离。此外，探讨了离子淌度-质谱（IM-MS）谱图分析时存在的多聚体碎片离子干扰问题。二级质谱使用注射泵直接进样，研究寡糖金属加合离子的裂解方式用于辨别同分异构体。最后，计算了两个寡糖系列的平均碰撞横截面积，可据此使用线性拟合预测同系列更长糖链离子的平均碰撞横截面积值。研究结果表明，漂移时间离子淌度-四极杆-飞行时间串联质谱法能有效实现寡糖同分异构体的定性分析。