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Jonathan I Amster - One of the best experts on this subject based on the ideXlab platform.
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electron detachment dissociation of synthetic heparan sulfate glycosaminoglycan tetrasaccharides varying in degree of sulfation and Hexuronic Acid stereochemistry
International Journal of Mass Spectrometry, 2012Co-Authors: Franklin E Leach, Sailaja Arungundram, Kanar Almafraji, Andre Venot, Geert-jan Boons, Jonathan I AmsterAbstract:Glycosaminoglycan (GAG) carbohydrates provide a challenging analytical target for structural determination due to their polydisperse nature, non-template biosynthesis, and labile sulfate modifications. The resultant structures, although heterogeneous, contain domains which indicate a sulfation pattern or code that correlates to specific function. Mass spectrometry, in particular electron detachment dissociation Fourier transform ion cyclotron resonance (EDD FT-ICR MS), provides a highly sensitive platform for GAG structural analysis by providing cross-ring cleavages for sulfation location and product ions specific to Hexuronic Acid stereochemistry. To investigate the effect of sulfation pattern and variations in stereochemistry on EDD spectra, a series of synthetic heparan sulfate (HS) tetrasaccharides are examined. Whereas previous studies have focused on lowly sulfated compounds (0.5–1 sulfate groups per disaccharide), the current work extends the application of EDD to more highly sulfated tetrasaccharides (1–2 sulfate groups per disaccharide) and presents the first EDD of a tetrasaccharide containing a sulfated Hexuronic Acid. For these more highly sulfated HS oligomers, alternative strategies are shown to be effective for extracting full structural details. These strategies inlcude sodium cation replacement of protons, for determining the sites of sulfation, and desulfation of the oligosaccharides for the generation of product ions for assigning uronic Acid stereochemistry.
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electron detachment dissociation of synthetic heparan sulfate glycosaminoglycan tetrasaccharides varying in degree of sulfation and Hexuronic Acid stereochemistry
International Journal of Mass Spectrometry, 2012Co-Authors: Franklin E Leach, Sailaja Arungundram, Kanar Almafraji, Andre Venot, Geert-jan Boons, Jonathan I AmsterAbstract:Glycosaminoglycan (GAG) carbohydrates provide a challenging analytical target for structural determination due to their polydisperse nature, non-template biosynthesis, and labile sulfate modifications. The resultant structures, although heterogeneous, contain domains which indicate a sulfation pattern or code that correlates to specific function. Mass spectrometry, in particular electron detachment dissociation Fourier transform ion cyclotron resonance (EDD FT-ICR MS), provides a highly sensitive platform for GAG structural analysis by providing cross-ring cleavages for sulfation location and product ions specific to Hexuronic Acid stereochemistry. To investigate the effect of sulfation pattern and variations in stereochemistry on EDD spectra, a series of synthetic heparan sulfate (HS) tetrasaccharides are examined. Whereas previous studies have focused on lowly sulfated compounds (0.5–1 sulfate groups per disaccharide), the current work extends the application of EDD to more highly sulfated tetrasaccharides (1–2 sulfate groups per disaccharide) and presents the first EDD of a tetrasaccharide containing a sulfated Hexuronic Acid. For these more highly sulfated HS oligomers, alternative strategies are shown to be effective for extracting full structural details. These strategies inlcude sodium cation replacement of protons, for determining the sites of sulfation, and desulfation of the oligosaccharides for the generation of product ions for assigning uronic Acid stereochemistry.
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Hexuronic Acid stereochemistry determination in chondroitin sulfate glycosaminoglycan oligosaccharides by electron detachment dissociation
Journal of the American Society for Mass Spectrometry, 2012Co-Authors: Franklin E Leach, Robert J. Linhardt, Mellisa Ly, Tatiana N Laremore, Jens J Wolff, Jacob Perlow, Jonathan I AmsterAbstract:Electron detachment dissociation (EDD) has previously provided stereo-specific product ions that allow for the assignment of the Acidic C-5stereochemistry in heparan sulfate glycosaminoglycans (GAGs), but application of the same methodology to an epimer pair in the chondroitin sulfate glycoform class does not provide the same result. A series of experiments have been conducted in which glycosaminoglycan precursor ions are independently activated by electron detachment dissociation (EDD), electron induced dissociation (EID), and negative electron transfer dissociation (NETD) to assign the stereochemistry in chondroitin sulfate (CS) epimers and investigate the mechanisms for product ion formation during EDD in CS glycoforms. This approach allows for the assignment of electronic excitation products formed by EID and detachment products to radical pathways in NETD, both of which occur simultaneously during EDD. The uronic Acid stereochemistry in electron detachment spectra produces intensity differences when assigned glycosidic and cross-ring cleavages are compared. The variations in the intensities of the doubly deprotonated 0,2X3 and Y3 ions have been shown to be indicative of CS-A/DS composition during the CID of binary mixtures. These ions can provide insight into the uronic Acid composition of binary mixtures in EDD, but the relative abundances, although reproducible, are low compared with those in a CID spectrum acquired on an ion trap. The application of principal component analysis (PCA) presents a multivariate approach to determining the uronic Acid stereochemistry spectra of these GAGs by taking advantage of the reproducible peak distributions produced by electron detachment.
Franklin E Leach - One of the best experts on this subject based on the ideXlab platform.
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electron detachment dissociation of synthetic heparan sulfate glycosaminoglycan tetrasaccharides varying in degree of sulfation and Hexuronic Acid stereochemistry
International Journal of Mass Spectrometry, 2012Co-Authors: Franklin E Leach, Sailaja Arungundram, Kanar Almafraji, Andre Venot, Geert-jan Boons, Jonathan I AmsterAbstract:Glycosaminoglycan (GAG) carbohydrates provide a challenging analytical target for structural determination due to their polydisperse nature, non-template biosynthesis, and labile sulfate modifications. The resultant structures, although heterogeneous, contain domains which indicate a sulfation pattern or code that correlates to specific function. Mass spectrometry, in particular electron detachment dissociation Fourier transform ion cyclotron resonance (EDD FT-ICR MS), provides a highly sensitive platform for GAG structural analysis by providing cross-ring cleavages for sulfation location and product ions specific to Hexuronic Acid stereochemistry. To investigate the effect of sulfation pattern and variations in stereochemistry on EDD spectra, a series of synthetic heparan sulfate (HS) tetrasaccharides are examined. Whereas previous studies have focused on lowly sulfated compounds (0.5–1 sulfate groups per disaccharide), the current work extends the application of EDD to more highly sulfated tetrasaccharides (1–2 sulfate groups per disaccharide) and presents the first EDD of a tetrasaccharide containing a sulfated Hexuronic Acid. For these more highly sulfated HS oligomers, alternative strategies are shown to be effective for extracting full structural details. These strategies inlcude sodium cation replacement of protons, for determining the sites of sulfation, and desulfation of the oligosaccharides for the generation of product ions for assigning uronic Acid stereochemistry.
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electron detachment dissociation of synthetic heparan sulfate glycosaminoglycan tetrasaccharides varying in degree of sulfation and Hexuronic Acid stereochemistry
International Journal of Mass Spectrometry, 2012Co-Authors: Franklin E Leach, Sailaja Arungundram, Kanar Almafraji, Andre Venot, Geert-jan Boons, Jonathan I AmsterAbstract:Glycosaminoglycan (GAG) carbohydrates provide a challenging analytical target for structural determination due to their polydisperse nature, non-template biosynthesis, and labile sulfate modifications. The resultant structures, although heterogeneous, contain domains which indicate a sulfation pattern or code that correlates to specific function. Mass spectrometry, in particular electron detachment dissociation Fourier transform ion cyclotron resonance (EDD FT-ICR MS), provides a highly sensitive platform for GAG structural analysis by providing cross-ring cleavages for sulfation location and product ions specific to Hexuronic Acid stereochemistry. To investigate the effect of sulfation pattern and variations in stereochemistry on EDD spectra, a series of synthetic heparan sulfate (HS) tetrasaccharides are examined. Whereas previous studies have focused on lowly sulfated compounds (0.5–1 sulfate groups per disaccharide), the current work extends the application of EDD to more highly sulfated tetrasaccharides (1–2 sulfate groups per disaccharide) and presents the first EDD of a tetrasaccharide containing a sulfated Hexuronic Acid. For these more highly sulfated HS oligomers, alternative strategies are shown to be effective for extracting full structural details. These strategies inlcude sodium cation replacement of protons, for determining the sites of sulfation, and desulfation of the oligosaccharides for the generation of product ions for assigning uronic Acid stereochemistry.
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Hexuronic Acid stereochemistry determination in chondroitin sulfate glycosaminoglycan oligosaccharides by electron detachment dissociation
Journal of the American Society for Mass Spectrometry, 2012Co-Authors: Franklin E Leach, Robert J. Linhardt, Mellisa Ly, Tatiana N Laremore, Jens J Wolff, Jacob Perlow, Jonathan I AmsterAbstract:Electron detachment dissociation (EDD) has previously provided stereo-specific product ions that allow for the assignment of the Acidic C-5stereochemistry in heparan sulfate glycosaminoglycans (GAGs), but application of the same methodology to an epimer pair in the chondroitin sulfate glycoform class does not provide the same result. A series of experiments have been conducted in which glycosaminoglycan precursor ions are independently activated by electron detachment dissociation (EDD), electron induced dissociation (EID), and negative electron transfer dissociation (NETD) to assign the stereochemistry in chondroitin sulfate (CS) epimers and investigate the mechanisms for product ion formation during EDD in CS glycoforms. This approach allows for the assignment of electronic excitation products formed by EID and detachment products to radical pathways in NETD, both of which occur simultaneously during EDD. The uronic Acid stereochemistry in electron detachment spectra produces intensity differences when assigned glycosidic and cross-ring cleavages are compared. The variations in the intensities of the doubly deprotonated 0,2X3 and Y3 ions have been shown to be indicative of CS-A/DS composition during the CID of binary mixtures. These ions can provide insight into the uronic Acid composition of binary mixtures in EDD, but the relative abundances, although reproducible, are low compared with those in a CID spectrum acquired on an ion trap. The application of principal component analysis (PCA) presents a multivariate approach to determining the uronic Acid stereochemistry spectra of these GAGs by taking advantage of the reproducible peak distributions produced by electron detachment.
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Multivariate analysis of electron detachment dissociation and infrared multiphoton dissociation mass spectra of heparan sulfate tetrasaccharides differing only in Hexuronic Acid stereochemistry.
Journal of the American Society for Mass Spectrometry, 2011Co-Authors: Han Bin Oh, Franklin E Leach, Sailaja Arungundram, Andre Venot, Geert-jan Boons, Kanar Al-mafraji, I. Jonathan AmsterAbstract:The structural characterization of glycosaminoglycan (GAG) carbohydrates by mass spectrometry has been a long-standing analytical challenge due to the inherent heterogeneity of these biomolecules, specifically polydispersity, variability in sulfation, and Hexuronic Acid stereochemistry. Recent advances in tandem mass spectrometry methods employing threshold and electron-based ion activation have resulted in the ability to determine the location of the labile sulfate modification as well as assign the stereochemistry of Hexuronic Acid residues. To facilitate the analysis of complex electron detachment dissociation (EDD) spectra, principal component analysis (PCA) is employed to differentiate the Hexuronic Acid stereochemistry of four synthetic GAG epimers whose EDD spectra are nearly identical upon visual inspection. For comparison, PCA is also applied to infrared multiphoton dissociation spectra (IRMPD) of the examined epimers. To assess the applicability of multivariate methods in GAG mixture analysis, PCA is utilized to identify the relative content of two epimers in a binary mixture.
Marion Kuschegullberg - One of the best experts on this subject based on the ideXlab platform.
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target selection of heparan sulfate Hexuronic Acid 2 o sulfotransferase
Glycobiology, 2010Co-Authors: Emanuel Smeds, Almir Feta, Marion KuschegullbergAbstract:The signaling of various molecules involved in development and regulation of cell growth are regulated by heparan sulfate (HS). Specific binding of HS to ligand proteins depends on the HS sulfati ...
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substrate specificity of the heparan sulfate Hexuronic Acid 2 o sulfotransferase
Biochemistry, 2001Co-Authors: Jianhui Rong, Ulf Lindahl, Hiroko Habuchi, Koji Kimata, Marion KuschegullbergAbstract:The interaction of heparan sulfate with different ligand proteins depends on the precise location of O-sulfate groups in the polysaccharide chain. We have previously shown that overexpression in human kidney 293 cells of a mouse mastocytoma 2-O-sulfotransferase (2-OST), previously thought to catalyze the transfer of sulfate from 3'-phosphoadenosine 5'-phosphosulfate to C2 of L-iduronyl residues, preferentially increases the level of 2-O-sulfation of D-glucuronyl units [Rong, J., Habuchi, H., Kimata, K., Lindahl, U., and Kusche-Gullberg, M. (2000) Biochem. J. 346, 463-468]. In the study presented here, we further investigated the substrate specificity of the mouse mastocytoma 2-OST. Different polysaccharide acceptor substrates were incubated with cell extracts from 2-OST-transfected 293 cells together with the sulfate donor 3'-phosphoadenosine 5'-phospho[(35)S]sulfate. Incubations with O-desulfated heparin, predominantly composed of [(4)alphaIdoA(1)-(4)alphaGlcNSO(3)(1)-](n)(), resulted in 2-O-sulfation of iduronic Acid. When, on the other hand, an N-sulfated capsular polysaccharide from Escherichia coli K5, with the structure [(4)betaGlcA(1)-(4)alphaGlcNSO(3)(1)-](n)(), was used as an acceptor, sulfate was transferred almost exclusively to C2 of glucuronic Acid. Substrates containing both iduronic and glucuronic Acid residues in about equal proportions strongly favored sulfation of iduronic Acid. In agreement with these results, the 2-OST was found to have a approximately 5-fold higher affinity for iduronic Acid-containing substrate disaccharide units (K(m) approximately 3.7 microM) than for glucuronic Acid-containing substrate disaccharide units (K(m) approximately 19.3 microM).
Ulf Lindahl - One of the best experts on this subject based on the ideXlab platform.
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pathophysiology of heparan sulphate many diseases few drugs
Journal of Internal Medicine, 2013Co-Authors: Ulf Lindahl, Lena KjellénAbstract:Heparan sulphate (HS) polysaccharides are covalently attached to the core proteins of various proteoglycans at cell surfaces and in the extracellular matrix. They are composed of alternating units of Hexuronic Acid and glucosamine, with sulphate substituents in complex and variable yet cell-specific patterns. Whereas HS is produced by virtually all cells in the body, heparin, a highly sulphated HS variant, is confined to connective-tissue-type mast cells. The polysaccharides interact with a multitude of proteins, mainly through ionic binding, and thereby control key processes in development and homoeostasis. Similar interactions also implicate HS in various pathophysiological settings, including cancer, amyloid diseases, infectious diseases, inflammatory conditions and some developmental disorders. Prospects for the development of HS-based drugs, which are still largely unrealized, are discussed.
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D-Glucuronyl C5-Epimerase in Heparin/Heparan Sulfate Biosynthesis
Handbook of Glycosyltransferases and Related Genes, 2002Co-Authors: Ulf LindahlAbstract:Glucuronyl C5 epimerase is a key enzyme in the biosynthesis of heparin and heparan sulfate (HS), which are complex sulfated glycosaminoglycans (GAGs) composed of alternating Hexuronic Acid and glucosamine residues. Both species are synthesized as proteoglycans but on different core proteins (Esko 1991). Heparin is produced by connective tissue-type mast cells, whereas HS is generated by most other mammalian (and many nonmammalian) cells. The process is initiated by glycosylation reactions that generate saccharide sequences composed of alternating D-glucuronic Acid (GlcA) and N-acetyl-D-glucosamine (GlcNAc) units, covalently bound to the respective core protein. The resulting polymer of (GlcAβ1,4-GlcNAcα1,4)n disaccharide repeats is modified through a series of reactions that includes N-deacetylation/N-sulfation of GlcNAc residues, C5-epimerization of the GlcA units to yield L-iduronic Acid (IdoA) residues, and finally O-sulfation at various positions (Lindahl et al. 1998). The modification reactions occur in a stepwise manner, such that early steps provide the substrates for subsequent reactions. Heparin is extensively sulfated and has a high content of IdoA (typically 50%–90% of the total Hexuronic Acid), whereas HS has a more varied structure, generally less sulfated than heparin and with lower IdoA content (typically 30%–55% of the total Hexuronic Acid) (Taylor et al. 1973). The corresponding enzymes have all recently been cloned (see also Chapter 67–69, 70–72).
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substrate specificity of the heparan sulfate Hexuronic Acid 2 o sulfotransferase
Biochemistry, 2001Co-Authors: Jianhui Rong, Ulf Lindahl, Hiroko Habuchi, Koji Kimata, Marion KuschegullbergAbstract:The interaction of heparan sulfate with different ligand proteins depends on the precise location of O-sulfate groups in the polysaccharide chain. We have previously shown that overexpression in human kidney 293 cells of a mouse mastocytoma 2-O-sulfotransferase (2-OST), previously thought to catalyze the transfer of sulfate from 3'-phosphoadenosine 5'-phosphosulfate to C2 of L-iduronyl residues, preferentially increases the level of 2-O-sulfation of D-glucuronyl units [Rong, J., Habuchi, H., Kimata, K., Lindahl, U., and Kusche-Gullberg, M. (2000) Biochem. J. 346, 463-468]. In the study presented here, we further investigated the substrate specificity of the mouse mastocytoma 2-OST. Different polysaccharide acceptor substrates were incubated with cell extracts from 2-OST-transfected 293 cells together with the sulfate donor 3'-phosphoadenosine 5'-phospho[(35)S]sulfate. Incubations with O-desulfated heparin, predominantly composed of [(4)alphaIdoA(1)-(4)alphaGlcNSO(3)(1)-](n)(), resulted in 2-O-sulfation of iduronic Acid. When, on the other hand, an N-sulfated capsular polysaccharide from Escherichia coli K5, with the structure [(4)betaGlcA(1)-(4)alphaGlcNSO(3)(1)-](n)(), was used as an acceptor, sulfate was transferred almost exclusively to C2 of glucuronic Acid. Substrates containing both iduronic and glucuronic Acid residues in about equal proportions strongly favored sulfation of iduronic Acid. In agreement with these results, the 2-OST was found to have a approximately 5-fold higher affinity for iduronic Acid-containing substrate disaccharide units (K(m) approximately 3.7 microM) than for glucuronic Acid-containing substrate disaccharide units (K(m) approximately 19.3 microM).
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Minimal sequence in heparin/heparan sulfate required for binding of basic fibroblast growth factor.
The Journal of biological chemistry, 1993Co-Authors: Marco Maccarana, B Casu, Ulf LindahlAbstract:Abstract Experiments based on interaction in free solution between basic fibroblast growth factor (FGF-2) and saccharides related to heparin/heparan sulfate showed that the growth factor binds to heparin and to selectively glucosaminyl 6-O-desulfated heparin but poorly to iduronosyl 2-O-desulfated heparin. 2-O-sulfate groups thus are essential to the interaction, whereas 6-O-sulfates are not required nor do they interfere with FGF-2 binding. Comparison of various bound/nonbound oligosaccharides implicated a minimal pentasaccharide sequence for FGF-2 binding, with the structure: -Hexuronic Acid-glucosamine N-sulfate-Hexuronic Acid-glucosamine N-sulfate-iduronic Acid 2-O-sulfate- (reducing terminus to the right). Such (overlapping) sequences are abundant in heparin, albeit heavily obscured by irrelevant O-sulfate groups, and occur also in heparan sulfate, with or without additional O-sulfates.
Geert-jan Boons - One of the best experts on this subject based on the ideXlab platform.
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Cryogenic Infrared Spectroscopy Reveals Structural Modularity in the Vibrational Fingerprints of Heparan Sulfate Diastereomers.
Analytical chemistry, 2020Co-Authors: Maike Lettow, Geert-jan Boons, Márkó Grabarics, Kim Greis, Eike Mucha, Daniel A. Thomas, Pradeep Chopra, Richard Karlsson, Jeremy E. Turnbull, Gerard MeijerAbstract:Heparan sulfate and heparin are highly Acidic polysaccharides with a linear sequence, consisting of alternating glucosamine and Hexuronic Acid building blocks. The identity of Hexuronic Acid units ...
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electron detachment dissociation of synthetic heparan sulfate glycosaminoglycan tetrasaccharides varying in degree of sulfation and Hexuronic Acid stereochemistry
International Journal of Mass Spectrometry, 2012Co-Authors: Franklin E Leach, Sailaja Arungundram, Kanar Almafraji, Andre Venot, Geert-jan Boons, Jonathan I AmsterAbstract:Glycosaminoglycan (GAG) carbohydrates provide a challenging analytical target for structural determination due to their polydisperse nature, non-template biosynthesis, and labile sulfate modifications. The resultant structures, although heterogeneous, contain domains which indicate a sulfation pattern or code that correlates to specific function. Mass spectrometry, in particular electron detachment dissociation Fourier transform ion cyclotron resonance (EDD FT-ICR MS), provides a highly sensitive platform for GAG structural analysis by providing cross-ring cleavages for sulfation location and product ions specific to Hexuronic Acid stereochemistry. To investigate the effect of sulfation pattern and variations in stereochemistry on EDD spectra, a series of synthetic heparan sulfate (HS) tetrasaccharides are examined. Whereas previous studies have focused on lowly sulfated compounds (0.5–1 sulfate groups per disaccharide), the current work extends the application of EDD to more highly sulfated tetrasaccharides (1–2 sulfate groups per disaccharide) and presents the first EDD of a tetrasaccharide containing a sulfated Hexuronic Acid. For these more highly sulfated HS oligomers, alternative strategies are shown to be effective for extracting full structural details. These strategies inlcude sodium cation replacement of protons, for determining the sites of sulfation, and desulfation of the oligosaccharides for the generation of product ions for assigning uronic Acid stereochemistry.
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electron detachment dissociation of synthetic heparan sulfate glycosaminoglycan tetrasaccharides varying in degree of sulfation and Hexuronic Acid stereochemistry
International Journal of Mass Spectrometry, 2012Co-Authors: Franklin E Leach, Sailaja Arungundram, Kanar Almafraji, Andre Venot, Geert-jan Boons, Jonathan I AmsterAbstract:Glycosaminoglycan (GAG) carbohydrates provide a challenging analytical target for structural determination due to their polydisperse nature, non-template biosynthesis, and labile sulfate modifications. The resultant structures, although heterogeneous, contain domains which indicate a sulfation pattern or code that correlates to specific function. Mass spectrometry, in particular electron detachment dissociation Fourier transform ion cyclotron resonance (EDD FT-ICR MS), provides a highly sensitive platform for GAG structural analysis by providing cross-ring cleavages for sulfation location and product ions specific to Hexuronic Acid stereochemistry. To investigate the effect of sulfation pattern and variations in stereochemistry on EDD spectra, a series of synthetic heparan sulfate (HS) tetrasaccharides are examined. Whereas previous studies have focused on lowly sulfated compounds (0.5–1 sulfate groups per disaccharide), the current work extends the application of EDD to more highly sulfated tetrasaccharides (1–2 sulfate groups per disaccharide) and presents the first EDD of a tetrasaccharide containing a sulfated Hexuronic Acid. For these more highly sulfated HS oligomers, alternative strategies are shown to be effective for extracting full structural details. These strategies inlcude sodium cation replacement of protons, for determining the sites of sulfation, and desulfation of the oligosaccharides for the generation of product ions for assigning uronic Acid stereochemistry.
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Multivariate analysis of electron detachment dissociation and infrared multiphoton dissociation mass spectra of heparan sulfate tetrasaccharides differing only in Hexuronic Acid stereochemistry.
Journal of the American Society for Mass Spectrometry, 2011Co-Authors: Han Bin Oh, Franklin E Leach, Sailaja Arungundram, Andre Venot, Geert-jan Boons, Kanar Al-mafraji, I. Jonathan AmsterAbstract:The structural characterization of glycosaminoglycan (GAG) carbohydrates by mass spectrometry has been a long-standing analytical challenge due to the inherent heterogeneity of these biomolecules, specifically polydispersity, variability in sulfation, and Hexuronic Acid stereochemistry. Recent advances in tandem mass spectrometry methods employing threshold and electron-based ion activation have resulted in the ability to determine the location of the labile sulfate modification as well as assign the stereochemistry of Hexuronic Acid residues. To facilitate the analysis of complex electron detachment dissociation (EDD) spectra, principal component analysis (PCA) is employed to differentiate the Hexuronic Acid stereochemistry of four synthetic GAG epimers whose EDD spectra are nearly identical upon visual inspection. For comparison, PCA is also applied to infrared multiphoton dissociation spectra (IRMPD) of the examined epimers. To assess the applicability of multivariate methods in GAG mixture analysis, PCA is utilized to identify the relative content of two epimers in a binary mixture.