Fig 1 - uploaded by Frederick L Kiechle
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The structure of ganglioside GM1 (panel A) and Svennerholm's nomenclature for gangliosides (panel B). G stands for ganglioside, M for monosialo-, D for disialo-, and T for trisialo-ganglioside. The number (n = 1, 2, 3, or 4) indicates the number of carbohydrate units and the sequence of migration of gangliosides on thin layer chromatograms.
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Glycosphingolipids are ubiquitous membrane constituents that are subdivided in neutral or acidic fractions (gangliosides and sulfatides). Their analysis requires extraction and separation by thin-layer chromatography or high-performance liquid chromatography. Ganglioside composition changes occur in response to variations in cellular morphology and...
Citations
... GSL comprise a vast group of remarkably heterogeneous biomolecules that are found in all eukaryotes, as well as some prokaryotes and viruses. GSL are ubiquitous membrane components, which are almost exclusively located on the outer leaflet of cell plasma membranes and in intracellular organelles (Fig. 1) [1,2]. GSL are also found in body fluids, where they either circulate freely or are transported in lipoproteins [3]. ...
... Neutral GSL (nGSL) include cerebrosides (1 glycan unit) and globosides (≥ 2 glycan units), while acidic GSL (aGSL) are subdivided into sialic acid-containing GSL called gangliosides and sulfated GSL called sulfatides with sulfate group at C3 hydroxyl of Gal. In contrast, basic GSL are rare [2]. ...
Glycosphingolipids (GSL) are a highly heterogeneous class of lipids representing the majority of the sphingolipid category. GSL are fundamental constituents of cellular membranes that have key roles in various biological processes, such as cellular signaling, recognition, and adhesion. Understanding the structural complexity of GSL is pivotal for unraveling their functional significance in a biological context, specifically their crucial role in the pathophysiology of various diseases. Mass spectrometry (MS) has emerged as a versatile and indispensable tool for the structural elucidation of GSL enabling a deeper understanding of their complex molecular structures and their key roles in cellular dynamics and patholophysiology. Here, we provide a thorough overview of MS techniques tailored for the analysis of GSL, emphasizing their utility in probing GSL intricate structures to advance our understanding of the functional relevance of GSL in health and disease. The application of tandem MS using diverse fragmentation techniques, including novel ion activation methodologies, in studying glycan sequences, linkage positions, and fatty acid composition is extensively discussed. Finally, we address current challenges, such as the detection of low-abundance species and the interpretation of complex spectra, and offer insights into potential solutions and future directions by improving MS instrumentation for enhanced sensitivity and resolution, developing novel ionization techniques, or integrating MS with other analytical approaches for comprehensive GSL characterization.
... Mohit Jaiswal and Trang T. Tran contribute equally to the current work. presentation patterns of glycans in the cell glycocalyx are associated with many diseases [2][3][4]. For example, more than 80% of cell surface glycoconjugates in the vertebrate brain are glycolipids [5]. ...
As new methods to interrogate glycan organization on cells develop, it is important to have molecular level understanding of how chemical fixation can impact results and interpretations. Site-directed spin labeling technologies are well suited to study how the spin label mobility is impacted by local environmental conditions, such as those imposed by cross-linking effects of paraformaldehyde cell fixation methods. Here, we utilize three different azide-containing sugars for metabolic glycan engineering with HeLa cells to incorporate azido glycans that are modified with a DBCO-based nitroxide moiety via click reaction. Continuous wave X-band electron paramagnetic resonance spectroscopy is employed to characterize how the chronological sequence of chemical fixation and spin labeling impacts the local mobility and accessibility of the nitroxide-labeled glycans in the glycocalyx of HeLa cells. Results demonstrate that chemical fixation with paraformaldehyde can alter local glycan mobility and care should be taken in the analysis of data in any study where chemical fixation and cellular labeling occur.
... Interestingly, compared with the C_O or MS_W groups, the level of GlcCer was significantly increased in the MS_W group. GlcCer is the simplest member and precursor of a fascinating class of membrane lipids, the glycosphingolipids, which are implicated in the pathogenesis of various diseases, including glycosphingolipidoses, peripheral neuropathies, and secretory diarrhea (Jeyakumar et al., ;van Meer et al., 2003;Zhang and Kiechle, 2004). The focus of our future attention will be on whether GlcCer plays a role in ameliorating depression-like behaviors and whether its role has brain area specificity. ...
The pathology of depression involves various factors including the interaction between genes and the environment. The deficiency of n-3 polyunsaturated fatty acids (n-3 PUFAs) in the brain and depressive symptoms are closely related. Krill oil contains abundant amounts of n-3 PUFAs incorporated in phosphatidylcholine. However, the effect of krill oil treatment on depression-like behaviors induced by chronic stress and its molecular mechanism in the brain remain poorly understood. Here, we used a chronic unpredictable mild stress (CUMS) model to evaluate the effect of krill oil on depression-like behaviors and explored its molecular mechanism through lipid metabolomics and mRNA profiles in the whole brain. We observed that CUMS-induced depression-like behaviors were ameliorated by krill oil supplementation in mice. The metabolism of glycerophospholipids and sphingolipids was disrupted by CUMS treatment, which were ameliorated after krill oil supplementation. Further analysis found that differently expressed genes after krill oil supplementation were mainly enriched in the membrane structures and neuroactive ligand–receptor interaction pathway, which may be responsible for the amelioration of CUMS-induced depression-like behaviors. Altogether, our results uncovered the relationship between lipid metabolism and CUMS, and provided new strategies for the prevention and treatment of depression.
... 24 GSLs are a subclass of glycolipids, consisting of carbohydrate moieties linked to the 1-hydroxyl group of a ceramide backbone via a β-linkage. 25 GSLs are ubiquitously embedded in the cell plasma membrane. The astonishing structural diversity of GSLs arises from linking hundreds of different glycan heads to tens of different ceramide chains, which are responsible for various biological activities, like regulation of cell growth, differentiation, and signaling. ...
... In the case of GalCer series GSLs, the precursor GalCer is transported to the Golgi complex where it can be sialylated to produce GM4 ganglioside or sulfated to produce sulfogalactolipids. 25 Glycosylphosphatidylinositol (GPI) often links to a glycan and acts as an anchor for a variety of cell surface proteins 31 as part of a post-translational protein modification process and is present in diverse eukaryotic species. 32,33 The GPI anchor consists of a phosphoethanolamine moiety linked to the terminal mannose of a highly conserved core glycan Manα1− 2Manα1−6Manα1−4GlcNH 2 α1−6myo-inositol, and the phospholipid tail (Figure 1). ...
Glycoconjugates are major constituents of mammalian cells that are formed via covalent conjugation of carbohydrates to other biomolecules like proteins and lipids and often expressed on the cell surfaces. Among the three major classes of glycoconjugates, proteoglycans and glycoproteins contain glycans linked to the protein backbone via amino acid residues such as Asn for N-linked glycans and Ser/Thr for O-linked glycans. In glycolipids, glycans are linked to a lipid component such as glycerol, polyisoprenyl pyrophosphate, fatty acid ester, or sphingolipid. Recently, glycoconjugates have become better structurally defined and biosynthetically understood, especially those associated with human diseases, and are accessible to new drug, diagnostic, and therapeutic developments. This review describes the status and new advances in the biological study and therapeutic applications of natural and synthetic glycoconjugates, including proteoglycans, glycoproteins, and glycolipids. The scope, limitations, and novel methodologies in the synthesis and clinical development of glycoconjugates including vaccines, glyco-remodeled antibodies, glycan-based adjuvants, glycan-specific receptor-mediated drug delivery platforms, etc., and their future prospectus are discussed.
... Glycosphingolipids (GSLs) are complex lipids consisting of glycans conjugated to a ceramide core and comprise a diverse group of over 300 molecules (Table 1) (D'Angelo et al., 2013;Nakayama et al., 2013). Although the nomenclature and classification of GSLs is complex (1978), they can be classified on the basis of their electrical charge as neutral, acidic (anionic), or basic (cationic), and according to their core structure as ganglio-series (N-acetylgalactosamine β-1,4 galactose β-1,4 glucose β-1,1′ Cer), globo-series (galactose α-1,4 galactose β-1,4 glucose 1,1′ Cer), isoglobo-series (galactose α-1,3 galactose β-1,4 glucose 1-1′ Cer), muco-series (galactose β-1,4 galactose β1-4 glucose 1,1′ Cer), lactoseries type 1 (galactose β-1,3-N-acetylglucosamine β-1,3 galactose β-1-4 glucose 1-1′ Cer), lactoseries type 2 (galactose β-1,4-N-acetylglucosamine β-1,3 galactose β-1,4 glucose 1-1′ Cer) and galaseries (galactose α-1,4 galactose 1-1′ Cer) (Keusch et al., 2000;Hakomori, 2003;Zhang and Kiechle, 2004;Kopitz, 2017). ...
At first glance, the biological function of globoside (Gb) clusters appears to be that of glycosphingolipid (GSL) receptors for bacterial toxins that mediate host-pathogen interaction. Indeed, certain bacterial toxin families have been evolutionarily arranged so that they can enter eukaryotic cells through GSL receptors. A closer look reveals this molecular arrangement allocated on a variety of eukaryotic cell membranes, with its role revolving around physiological regulation and pathological processes. What makes Gb such a ubiquitous functional arrangement? Perhaps its peculiarity is underpinned by the molecular structure itself, the nature of Gb-bound ligands, or the intracellular trafficking unleashed by those ligands. Moreover, Gb biological conspicuousness may not lie on intrinsic properties or on its enzymatic synthesis/degradation pathways. The present review traverses these biological aspects, focusing mainly on globotriaosylceramide (Gb3), a GSL molecule present in cell membranes of distinct cell types, and proposes a wrap-up discussion with a phylogenetic view and the physiological and pathological functional alternatives.
... Sphingolipids and gangliosides are metabolically interconnected structural and signalling components of cell membranes, and deregulation in their metabolism is linked to key human diseases including cancer (1,2). Ceramides are the central hub of the sphingolipid pathway, and modifications at their 3'hydroxyl terminal lead to structurally diverse classes of sphingolipids like glucosylceramides, sphingomyelins, and ceramide-1-phosphates with a distinct role in different facets of tumorigenesis ( Figure 1A) (3). ...
Sphingolipid and ganglioside metabolic pathways are crucial components of cell signalling, having established roles in tumor cell proliferation, invasion, and migration. However, regulatory mechanisms controlling sphingolipid and ganglioside synthesis in mammalian cells is less known. Here, we show that RICTOR, the regulatory subunit of mTORC2, regulates the synthesis of sphingolipids and gangliosides in Luminal breast cancer-specific MCF-7 cells through transcriptional and epigenetic mechanisms. RICTOR regulates glucosylceramide levels by modulating the expression of UDP-Glucose Ceramide Glucosyl transferase (UGCG). We identify Zinc Finger protein X-linked (ZFX) as a RICTOR-responsive transcription factor whose recruitment to the UGCG promoter is regulated by DNA methyltransferases and histone demethylase (KDM5A) that are known AKT substrates. We further demonstrate that RICTOR regulates the synthesis of GD3 gangliosides through ZFX and UGCG, and triggers the activation of the EGFR signalling pathway, thereby promoting tumor growth. In line with our findings in cell culture and mice models, we observe an elevated expression of RICTOR, ZFX, and UGCG in Indian Luminal breast cancer patient samples, and in TCGA and METABRIC datasets. Together, we establish a key regulatory circuit, RICTOR-AKT-ZFX-UGCG-Ganglioside-EGFR-AKT, and elucidate its contribution to breast cancer progression.
... Embedded in the outer leaflet of the plasma membrane, the ceramide lipid tail consisting of long-chain bases (sphingosine) and fatty acyl chains of different lengths and degrees of saturation [4,5] is glycosidically linked to a glycan headgroup containing one or more sialic acid residues. Extended into the extracellular environment, the glycan core, especially through the terminal sialic acid residues [6], participates in specific and essential biological functions of the brain [3], such as cell-to-cell recognition/communication and signaling, modulating, or triggering a variety of biological events, including those related to brain development, maturation, and aging [7][8][9][10]. GGs are also strongly correlated with brain disorders through aberrant glycosylation pathways. ...
Gangliosides are effective biochemical markers of brain pathologies, being also in the focus of research as potential therapeutic targets. Accurate brain ganglioside mapping is an essential requirement for correlating the specificity of their composition with a certain pathological state and establishing a well-defined set of biomarkers. Among all bioanalytical methods conceived for this purpose, mass spectrometry (MS) has developed into one of the most valuable, due to the wealth and consistency of structural information provided. In this context, the present article reviews the achievements of MS in discovery and structural analysis of gangliosides associated with severe brain pathologies. The first part is dedicated to the contributions of MS in the assessment of ganglioside composition and role in the specific neurodegenerative disorders: Alzheimer's and Parkinson's diseases. A large subsequent section is devoted to cephalic disorders (CD), with an emphasis on the MS of gangliosides in anencephaly, the most common and severe disease in the CD spectrum. The last part is focused on the major accomplishments of MS-based methods in the discovery of ganglioside species, which are associated with primary and secondary brain tumors and may either facilitate an early diagnosis or represent target molecules for immunotherapy oriented against brain cancers.
... Glycosphingolipids are ubiquitous membrane components that contribute to the pathogenesis of various diseases, including glycosphingolipidosis, anti-ganglioside antibody-induced peripheral neuropathies, and secretory diarrhea (Kolter, 2011). Gangliosides (a group of glycosphingolipids) are a common component of vertebrate cells and play an important role in the pathology of various human diseases; they are also potential diagnostic markers and therapeutic targets for cancer (Zhang and Kiechle, 2004). As a result, there is a great demand for them, and considerable work being done to synthesize these substances (Vankar and Schmidt, 2000). ...
Until recently, glycosidases, naturally hydrolyzing carbohydrate-active enzymes, have found few synthetic applications in industry, being primarily used for cleaving unwanted carbohydrates. With the establishment of glycosynthase and transglycosidase technology by genetic engineering, the view of glycosidases as industrial biotechnology tools has started to change. Their easy production, affordability, robustness, and substrate versatility, added to the possibility of controlling undesired side hydrolysis by enzyme engineering, have made glycosidases competitive synthetic tools. Current promising applications of engineered glycosidases include the production of well-defined chitooligomers, precious galactooligosaccharides or specialty chemicals such as glycosylated flavonoids. Other synthetic pathways leading to human milk oligosaccharides or remodeled antibodies are on the horizon. This work provides an overview of the synthetic achievements to date for glycosidases, emphasizing the latest trends and outlining possible developments in the field.
... Glycosphingolipids are ubiquitous membrane components that contribute to the pathogenesis of various diseases, including glycosphingolipidosis, anti-ganglioside antibody-induced peripheral neuropathies, and secretory diarrhea (Kolter, 2011). Gangliosides (a group of glycosphingolipids) are a common component of vertebrate cells and play an important role in the pathology of various human diseases; they are also potential diagnostic markers and therapeutic targets for cancer (Zhang and Kiechle, 2004). As a result, there is a great demand for them, and considerable work being done to synthesize these substances (Vankar and Schmidt, 2000). ...
Until recently, glycosidases, naturally hydrolyzing carbohydrate-active enzymes, have found few synthetic applications in industry, being primarily used for cleaving unwanted carbohydrates. With the establishment of glycosynthase and transglycosidase technology by genetic engineering, the view of glycosidases as industrial biotechnology tools has started to change. Their easy production, affordability, robustness, and substrate versatility, added to the possibility of controlling undesired side hydrolysis by enzyme engineering, have made glycosidases competitive synthetic tools. Current promising applications of engineered glycosidases include the production of well-defined chitooligomers, precious galactooligosaccharides or specialty chemicals such as glycosylated flavonoids. Other synthetic pathways leading to human milk oligosaccharides or remodeled antibodies are on the horizon. This work provides an overview of the synthetic achievements to date for glycosidases, emphasizing the latest trends and outlining possible developments in the field.
... Glycosphingolipids (GSL) are ubiquitous membrane components [1] predominantly found in the outer leaflet of the cell plasma membrane [2][3][4] of virtually all eukaryotic species [5] and some bacteria [6], where they are included in lipid rafts [1,5] together with cholesterol clusters, which serves as binding sites or receptors [4]. Furthermore, they have been found in membranes of intracellular organelles [3], and they also circulate in serum either in free or protein-bounded form [7]. ...
... Glycosphingolipids (GSL) are ubiquitous membrane components [1] predominantly found in the outer leaflet of the cell plasma membrane [2][3][4] of virtually all eukaryotic species [5] and some bacteria [6], where they are included in lipid rafts [1,5] together with cholesterol clusters, which serves as binding sites or receptors [4]. Furthermore, they have been found in membranes of intracellular organelles [3], and they also circulate in serum either in free or protein-bounded form [7]. ...
... Furthermore, they have been found in membranes of intracellular organelles [3], and they also circulate in serum either in free or protein-bounded form [7]. These amphipathic compounds represent one of the major and most structurally heterogeneous subclass of sphingolipids [8,9] comprised of a hydrophilic head group with at least one monosaccharide residue attached via glycosidic linkage to a hydrophobic ceramide moiety [1,2,4,10]. GSL are immensely complex due to multiple variations in composition, binding positions, branching [2,11] and modifications (e.g., sulfation, sialylation) of a saccharide core [8] as well as in ceramide backbone (e.g., hydroxylation) [12]. ...
Glycosphingolipids (GSL) represent a highly heterogeneous class of lipids with many cellular functions, implicated in a wide spectrum of human diseases. Their isolation, detection, and comprehensive structural analysis is a challenging task due to the structural diversity of GSL molecules. In this work, GSL subclasses are isolated from human plasma using an optimized monophasic ethanol–water solvent system capable to recover a broad range of GSL species. Obtained deproteinized plasma is subsequently purified and concentrated by C18-based solid-phase extraction (SPE). The hydrophilic interaction liquid chromatography coupled to electrospray ionization linear ion trap tandem mass spectrometry (HILIC-ESI-LIT-MS/MS) is used for GSL analysis in the human plasma extract. Our results provide an in-depth profiling and structural characterization of glycosphingolipid and some phospholipid subclasses identified in the human plasma based on their retention times and the interpretation of tandem mass spectra. The structural composition of particular lipid species is readily characterized based on the detailed interpretation of mass spectrometry (MS) and tandem mass spectrometry (MS/MS) spectra and further confirmed by specific fragmentation behavior following predictable patterns, which yields to the unambiguous identification of 154 GSL species within 7 lipid subclasses and 77 phospholipids representing the highest number of GSL species ever reported in the human plasma. The developed HILIC-ESI-MS/MS method can be used for further clinical and biological research of GSL in the human blood or other biological samples.
