Cleavage of the antithrombin III binding site in heparin by heparinases and its implication in the generation of low molecular weight heparin

Shriver, Zachary; Sundaram, Mallikarjun; Venkataraman, Ganesh; Fareed, Jawed; Linhardt, Robert J.; Biemann, K.; Sasisekharan, Ram

doi:10.1073/pnas.97.19.10365

Cited by 122 publications

(114 citation statements)

References 21 publications

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“…HLGAG oligosaccharides may be analyzed by negative ESI-MS yielding abundant deprotonated ions and less abundant sodium, potassium, and ammonium adducts [10,[13][14][15]. Solvent conditions and skimmer voltages may be adjusted to minimize the spectral complexity that results from the occurrence of ions formed via loss(es) of SO 3 in the ion source, to disfavor adduct formation and to control charge state formation.…”

Section: Esi Conditions For Production Of Highly Charged Ions From Sumentioning

confidence: 99%

“…The abundances of the charge states observed under each condition are dictated by factors including the molecular conformation of the ion in solution 14 , instrumental ion source conditions [15], pH [16], solvent polarity [17], and solvent volatility [18,19]. In these experiments the pH and instrument operating conditions were held constant.…”

Section: Esi Conditions For Production Of Highly Charged Ions From Sumentioning

confidence: 99%

“…HLGAGs have been paired with basic peptides [14,15,20] and ammonium counterions [10,25,26] to facilitate mass spectrometric analysis. Ion pairing reduces the abundances of ions that show sodium adduction, prevents losses of SO 3 , yields one dominant charge state, and thereby produces a simple mass spectrum [21].…”

Section: Esi Conditions For Production Of Highly Charged Ions From Sumentioning

confidence: 99%

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Competing fragmentation processes in tandem mass spectra of heparin-like glycosaminoglycans

Naggar

Costello

Zaia

2004

J. Am. Soc. Mass Spectrom.

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Heparin-like glycosaminoglycans (HLGAGs) are highly sulfated, linear carbohydrates attached to proteoglycan core proteins and expressed on cell surfaces and in basement membranes. These carbohydrates bind several families of growth factors and growth factor receptors and act as coreceptors for these molecules. Tandem mass spectrometry has the potential to increase our understanding of the biological significance of HLGAG expression by providing a facile means for sequencing these molecules without the need for time-consuming total purification. The challenge for tandem mass spectrometric analysis of HLGAGs is to produce abundant ions derived via glycosidic bond cleavages while minimizing the abundances of ions produced from elimination of the fragile sulfate groups. This work describes the competing fragmentation pathways that result from dissociation of high negative charge state ions generated from HLGAGs. Glycosidic bond cleavage ion formation competes with losses of equivalents of H 2 SO 4 , resulting in complex ion patterns. For the most highly sulfated structure examined, an octasulfated tetramer, an unusual loss of charge from the precursor ion was observed, accompanied by low abundance ions originating from subsequent backbone cleavages. These results demonstrate that fragmentation processes competing with glycosidic bond cleavages are more favored for highly sulfated HLGAG ions. In conclusion, reduction of charge-charge repulsions, such as is achieved by pairing the HLGAG ions with metal cations, is necessary in order to minimize the abundances of ions derived via fragmentation processes that compete with glycosidic bond cleavages. Sequence determination, including localization of sulfate groups on HLGAGs, is of significant interest, as their biological activities are believed to be expressed through patterns of sulfation and uronic acid epimerization [4,5].HLGAGs have traditionally proven difficult to analyze using mass spectrometry. Efforts to analyze these molecules using fast atom bombardment MS resulted in the observation of losses of SO 3 and NaSO 3 from the precursor ion, in addition to glycosidic bond cleavage ions [6 -10]. The poor sensitivity, relative to that achieved for peptides of comparable size, and lack of an intact precursor ion meant that tandem MS was not a viable analysis option when FAB ionization was employed. Although HLGAGs produce weak signals by matrix-assisted laser desorption/ionization (MALDI), much more abundant ions are detected from complexes between these molecules and a basic peptide [11][12][13]. In conjunction with enzymatic and/or chemical degradation, this MALDI sample preparation technique has been effectively used to sequence highly purified HLGAGs [14 -17]. Because the ions are detected in the positive mode, however, the charge is carried by the basic peptide and tandem MS of the HLGAG is not possible.Because HLGAGs and other sulfated carbohydrates produce abundant negative ions using electrospray ionization (ESI) without requiring basic peptide complexation [18...

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Section: Esi Conditions For Production Of Highly Charged Ions From Sumentioning

confidence: 99%

Section: Esi Conditions For Production Of Highly Charged Ions From Sumentioning

confidence: 99%

Section: Esi Conditions For Production Of Highly Charged Ions From Sumentioning

confidence: 99%

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Competing fragmentation processes in tandem mass spectra of heparin-like glycosaminoglycans

Naggar

Costello

Zaia

2004

J. Am. Soc. Mass Spectrom.

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“…MALDI time-of-flight (TOF) MS (19,26) and 23,33,34) have been the most favored approaches for the analysis of GAG-derived oligosaccharides. Both ESI-MS and MALDI-TOF-MS enjoy particular advantages in the analysis of large polar macromolecules.…”

Section: Liquid Chromatography/mass Spectrometry (Lc/ms)mentioning

confidence: 99%

Liquid Chromatography/Mass Spectrometry Sequencing Approach for Highly Sulfated Heparin-derived Oligosaccharides

Thanawiroon

Rice

Toida

et al. 2004

Journal of Biological Chemistry

146

152

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Liquid chromatography/mass spectrometry (LC/MS)is applied to the analysis of complex mixtures of oligosaccharides obtained through the controlled, heparinase-catalyzed depolymerization of heparin. Reversedphase ion-pairing chromatography, utilizing a volatile mobile phase, results in the high resolution separation of highly sulfated, heparin-derived oligosaccharides. Simultaneous detection by UV absorbance and electrospray ionization-mass spectrometry (ESI-MS) provides important structural information on the oligosaccharide components of this mixture. Highly sensitive and easily interpretable spectra were obtained through post-column addition of tributylamine in acetonitrile. High resolution mass spectrometry afforded elemental composition of many known and previously unknown heparin-derived oligosaccharides. UV in combination with MS detection led to the identification of oligosaccharides arising from the original non-reducing end (NRE) of the heparin chain. The structural identification of these oligosaccharides provided sequence from a reading frame that begins at the non-reducing terminus of the heparin chain. Interestingly, 16 NRE oligosaccharides are observed, having both an even and an odd number of saccharide residues, most of which are not predicted based on biosynthesis or known pathways of heparin catabolism. Quantification of these NRE oligosaccharides afforded a number-averaged molecular weight consistent with that expected for the pharmaceutical heparin used in this analysis. Molecular ions could be assigned for oligosaccharides as large as a tetradecasaccharide, having a mass of 4625 Da and a net charge of ؊32. Furthermore, MS detection was demonstrated for oligosaccharides with up to 30 saccharide units having a mass of >10,000 Da and a net charge of ؊60.The structural elucidation of complex carbohydrates remains one of the most difficult challenges for chemists, often requiring the application of multiple analytical approaches (1-5). Glycosaminoglycans (GAGs), 1 and heparin in particular, have proven to be extremely difficult to analyze because of high negative charge, polydispersity, and sequence heterogeneity (6, 7). Heparin and low molecular weight heparins, prepared through the controlled chemical or enzymatic fragmentation of heparin (8), are widely used as clinical anticoagulants. Despite their medical importance, these drugs are relatively uncharacterized in terms of their chemical structure. Moreover, heparin and the structurally related heparan sulfate exhibit many additional biological activities, making them of great interest in new drug discovery (9). Numerous challenges can arise from the structural investigation of biologically active heparin oligosaccharides, particularly those in recognition systems involving specific protein-carbohydrate interactions (10, 11). Such biologically important oligosaccharides often contain rare sequences (12, 13) and are present only in minute, often picomole quantities. New derivatization methods (14, 15), chromatography (15, 16), electrophoresis-ba...

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“…Demonstrated advantages of LMWH are the greater bioavailability at low doses, the longer half-life, and the more predictable dose response. In addition, LMWH are at least as safe and effective as unfractionated heparin in the treatment of established deep vein thrombosis, but have the advantage that they can be administered once or twice daily without laboratory monitoring [18].…”

Section: Khalife G Ghazeeri Open Journal Of Obstetrics and Gynecmentioning

confidence: 99%

Recurrent Implantation Failure and Low Molecular Weight Heparin

Khalife¹,

Ghazeeri²

2018

OJOG

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Implantation of the embryo into the endometrium is the first step in the establishment of pregnancy. This process is complex, and depends on many factors. Recurrent implantation failure is a source of distress to patients and specialists. It is defined as failure to achieve a viable pregnancy, following ">3 embryo transfers with high quality embryos or the transfer of ≥ 10 embryos in multiple transfers". Thrombophilic conditions that contribute to recurrent implantation failure are the main concern in this review. The mechanism of implantation failure is believed to be due to decreased blood flow to the endometrium and placenta which can hinder normal endometrial receptivity leading to miscarriage. Defects in early placentation resulting in pregnancy failure, have focused attention on the therapeutic potential of low molecular weight heparin in the implantation process. Heparin has a role at all stages of implantation to improve pregnancy outcomes. There are controversies in literature regarding the association between thrombophilia and recurrent implantation failure and available literature regarding this issue is very heterogeneous. Various investigators, have shown that women with RIF are more likely to have a thrombophilia disorder, yet a clear cause cannot be acknowledged from these studies. Heparin treatment has been evaluated in several studies, showing conflicting evidence. However, several studies have pointed out that it may play a role in a subset of patients who presents a thrombophilia mutation, thus the group of patients that might benefit is needed to be identified. This review is dedicated to evaluate the published literature about the role of low molecular weight heparin in case of recurrent implantation failure with or without the presence of thrombophilia.

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Cleavage of the antithrombin III binding site in heparin by heparinases and its implication in the generation of low molecular weight heparin

Cited by 122 publications

References 21 publications

Competing fragmentation processes in tandem mass spectra of heparin-like glycosaminoglycans

Competing fragmentation processes in tandem mass spectra of heparin-like glycosaminoglycans

Liquid Chromatography/Mass Spectrometry Sequencing Approach for Highly Sulfated Heparin-derived Oligosaccharides

Recurrent Implantation Failure and Low Molecular Weight Heparin

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