Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.26.9 (ribonuclease)
 6,589 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A polymeric, secondary amine stationary phase was used to develop a method for the separation of 2-aminobenzamide (AB)-labeled neutral and anionic oligosaccharides from glycoproteins. Sequential hydrophilic interaction liquid and anion exchange chromatography were performed in the same chromatographic analysis (multimode HPLC) and unambiguous separation between neutral and anionic oligosaccharides was accomplished. Improved resolution was also achieved. The oligomannosidic-type structures from bovine ribonuclease B (Man4GlcNAc-AB-Man9GlcNAc-AB) were separated not only by size, but also by the branch location of terminal Manalpha(1 --> 2)-linked residues. Sialylated lactosamine-type oligosaccharides from human alpha1 acid glycoprotein were resolved according to charge, Fuc content, and the number of Galbeta(1 --> 4)Galbeta(1 --> 3) repeats. The minor fucosylated and polylactosamine species were well separated from the major sialylated tetra-antennary oligosaccharides. Volatile mobile phases were used to minimize sample handling for matrix assisted laser desorption mass spectrometric analysis of peak fractions. Novel polylactosamine structures (3-5 repeats) from alpha1-acid glycoprotein were deduced from the molecular weight analysis. Multimode HPLC should prove useful for preparing low pmol quantities of fluorescently labeled oligosaccharides with fewer steps and minimal sample handling for mass spectrometric analysis, important requisites for structural studies of sample-limited glycoproteins.
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PMID:Multimode high-performance liquid chromatography of fluorescently labeled oligosaccharides from glycoproteins. 881 8

Oligomers of glucose and oligosaccharides released from glycoproteins were derivatized with 2-aminobenzamide. As this fluorophore imparts no charge to the oligosaccharides, several strategies were investigated to achieve capillary electrophoresis (CE) separation of both neutral and charged derivatized glycans. Micellar electrokinetic capillary chromatography (MEKC) with the addition of anionic surfactants was evaluated as a first approach: sodium dodecyl sulfate (SDS) produced the best separation of the oligoglucose fragments, where the migration was inversely related to their degree of polymerization. To demonstrate the applicability of this method for complex carbohydrate analysis, oligosaccharide mixtures derived from ribonuclease B (RNase B) and alpha-acid glycoprotein (alpha-AGP) were analyzed. A satisfactory separation for the high-mannose structures found in RNase B could be obtained, whereas charged oligosaccharides from alpha-AGP were poorly resolved. Cyclodextrin-modified CE was chosen as the second approach: the effect of the addition of sulfobutylether-beta-cyclodextrin (SBE-beta-CD) or sulfobutylether-gamma-cyclodextrin (SBE-gamma-CD) on the electrophoretic mobilities and resolution of neutral and charged oligosaccharides was then studied. Selectivity of sialylated structures could be further improved by using anionic cyclodextrins (CDs) instead of micelles. However, this latter approach failed to baseline-resolve the different high-mannose structures of RNase B. A successful separation of the complex mixture of oligosaccharides from alphaalpha-AGP was obtained with the addition of 4% of SBE-gamma-CD and triethylamine (TEA) in a phosphate buffer, pH 6.7.
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PMID:Investigation of micelles and anionic cyclodextrins as pseudostationary phases for the capillary electrophoresis separation of oligosaccharides derivatized with 2-amino-benzamide. 984 71

Ultraviolet photodissociation (UVPD) produces complementary fragmentation to collision-induced dissociation (CID) when implemented for activation of fluorescently labeled oligosaccharide and glycan ions. Reductive amination of oligosaccharides with fluorophore reagents results in efficient photon absorption at 355 nm, producing fragment ions from the nonreducing end that do not contain the appended fluorophore. In contrast to the fragment ions observed upon UVPD (A- and C-type ions), CID produces mainly reducing end fragments retaining the fluorophore (Y-type ions). UVPD affords better isomeric differentiation of both the lacto-N-fucopentaoses series and the lacto-N-difucohexaoses series, but in general, the combination of UVPD and CID offers the most diagnostic elucidation of complex branched oligosaccharides. Four fluorophores yielded similar MS/MS results; however, 6-aminoquinoline (6-AQ), 2-amino-9(10 H)-acridone (AMAC) and 7-aminomethylcoumarin (AMC) afforded more efficient photon absorption and subsequent dissociation than 2-aminobenzamide (2-AB). UVPD also was useful for characterization of glycans released from ribonuclease B and derivatized with 6-AQ. Lastly, electron photodetachment dissociation of oligosaccharides derivatized with 7-amino-1,3-naphthalenedisulfonic acid (AGA) yielded unique cross-ring cleavages similar to those obtained by electron detachment dissociation.
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PMID:Ultraviolet photodissociation at 355 nm of fluorescently labeled oligosaccharides. 1850 68

Tandem mass spectrometry of glycans and glycoconjugates in protonated form is known to result in rearrangement reactions leading to internal residue loss. Here we studied the occurrence of hexose rearrangements in tandem mass spectrometry of N-glycopeptides and reductively aminated N-glycans by MALDI-TOF/TOF-MS/MS and ESI-ion trap-MS/MS. Fragmentation of proton adducts of oligomannosidic N-glycans of ribonuclease B that were labeled with 2-aminobenzamide and 2-aminobenzoic acid resulted in transfer of one to five hexose residues to the fluorescently tagged innermost N-acetylglucosamine. Glycopeptides from various biological sources with oligomannosidic glycans were likewise shown to undergo hexose rearrangement reactions, resulting in chitobiose cleavage products that have acquired one or two hexose moieties. Tryptic immunoglobulin G Fc-glycopeptides with biantennary N-glycans likewise showed hexose rearrangements resulting in hexose transfer to the peptide moiety retaining the innermost N-acetylglucosamine. Thus, as a general phenomenon, tandem mass spectrometry of reductively aminated glycans as well as glycopeptides may result in hexose rearrangements. This characteristic of glycopeptide MS/MS has to be considered when developing tools for de novo glycopeptide structural analysis.
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PMID:Hexose rearrangements upon fragmentation of N-glycopeptides and reductively aminated N-glycans. 1941 47