Leishmania tropica: intraspecific polymorphisms in lipophosphoglycan correlate with transmission by different Phlebotomus species.
AuthorsSoares RP, Barron T, McCoy-Simandle K, Svobodova M, Warburg A, Turco SJ
JournalExp Parasitol
PubMed ID15208044
'Lipophosphoglycan (LPG) is a dominant surface molecule of Leishmania promastigotes which has been shown to be critical for parasite-sand fly vector interactions. To provide additional evidence for its importance in transmission, the LPGs from three Leishmania tropica strains that differ in their capability to infect sand flies, were biochemically characterized. ... More
Receptor for anionic pyrene derivatives provides the basis for new biomembrane assays.
AuthorsWinschel CA, Kalidindi A, Zgani I, Magruder JL, Sidorov V
JournalJ Am Chem Soc
PubMed ID16231924
'This study describes a new receptor cyclen 1 capable of strong selective binding of pyrene-based anionic dyes under near-physiological conditions. This receptor comprises four naphthylthiourea groups tethered to a cyclen core via an ester linkage. The complexation behavior of cyclen 1 receptor is characterized by a series of (1)H NMR, ... More
Profiling glycoprotein n-linked oligosaccharide by capillary electrophoresis.
AuthorsChen FT, Evangelista RA
JournalElectrophoresis
PubMed ID9848672
'A method for analysis of N-linked oligosaccharides derived from glycoproteins including sialic acid-containing species is presented. It is based on the combination of specific chemical and enzymatic conversions coupled with capillary electrophoretic (CE) separation and laser-induced fluorescence (LIF) detection. Glycoproteins were heat-denatured in the presence of a reducing agent and ... More
Analysis of neutral N-linked oligosaccharides from antibodies using free-solution capillary electrophoresis in bare fused-silica capillaries.
AuthorsPatrick JS, Rener BP, Clanton GS, Lagu AL
JournalMethods Mol Biol
PubMed ID15163856
'Conditions for the enzymatic release, chemical derivatization, and analysis of oligosaccharides from the consensus glycosylation sites on antibodies are described. Release of the oligosaccharides is from the native protein. The APTS derivatives of the released oligosaccharides are then analyzed by capillary electrophoresis (CE) using a free solution separation in a ... More
Analysis of chitin oligosaccharides by capillary electrophoresis with laser-induced fluorescence.
AuthorsWang CY, Hsieh YZ
JournalJ Chromatogr A
PubMed ID12498275
'A method, using capillary electrophoresis (CE) with laser-induced fluorescence (LIF) detection for analyzing chitin oligosaccharides is described. Chitin oligosaccharides were derivatized with 9-aminopyrene-1,4,6-trisulfonate (APTS) via reductive amination at 37 degrees C for 16 h (optimized conditions). The APTS-chitin oligosaccharides were analyzed using either an acidic citric acid-phosphate buffer or an ... More
Miniaturization in carbohydrate analysis.
AuthorsSuzuki S, Honda S
JournalElectrophoresis
PubMed ID14613182
'Recent progress of microchip electrophoresis (ME) of carbohydrates is overviewed. Carbohydrate analysis by ME encounters difficulties such as lack of electric charge and deficiency of a chromophore/fluorophore in analyte molecules, however, it benefits from the accumulated knowledge of capillary electrophoresis (CE) and rapid separation of simple sugars also by ME, ... More
Species-specific variation in glycosylation of IgG: evidence for the species-specific sialylation and branch-specific galactosylation and importance for engineering recombinant glycoprotein therapeutics.
AuthorsRaju TS, Briggs JB, Borge SM, Jones AJ
JournalGlycobiology
PubMed ID10764836
'Immunoglobulins (IgG) are soluble serum glycoproteins in which the oligosaccharides play significant roles in the bioactivity and pharmacokinetics. Recombinant immuno-globulins (rIgG) produced in different host cells by recombinant DNA technology are becoming major therapeutic agents to treat life threatening diseases such as cancer. Since glycosylation is cell type specific, rIgGs ... More
An essential oligomannosidic glycan chain in the catalytic domain of autotaxin, a secreted lysophospholipase-D.
AuthorsJansen S, Callewaert N, Dewerte I, Andries M, Ceulemans H, Bollen M
JournalJ Biol Chem
PubMed ID17307740
'Autotaxin/NPP2, a secreted lysophospholipase-D, promotes cell proliferation, survival, and motility by generating the signaling molecule lysophosphatidic acid. Here we show that ectonucleotide pyrophosphatase/phosphodiesterase 2 (NPP2) is N-glycosylated on Asn-53, Asn-410, and Asn-524. Mutagenesis and deglycosylation experiments revealed that only the glycosylation of Asn-524 is essential for the expression of the ... More
Carbohydrate analysis of a chimeric recombinant monoclonal antibody by capillary electrophoresis with laser-induced fluorescence detection.
AuthorsMa S, Nashabeh W
JournalAnal Chem
PubMed ID10575965
'A general method for the analysis of asparaginyl-linked (N-linked) carbohydrate moieties of an IgG1 monoclonal antibody is described here. The antibody, rituximab, is a mouse/human chimeric antibody to human CD20 antigen. The glycans present on rituximab are neutral complex biantennary oligosaccharides with zero, one, and two terminal galactose residues (G0, ... More
Characterization of mannooligosaccharide caps in mycobacterial lipoarabinomannan by capillary electrophoresis/electrospray mass spectrometry.
AuthorsMonsarrat B, Brando T, Condouret P, Nigou J, Puzo G
JournalGlycobiology
PubMed ID10089207
'A new analytical approach based on capillary electrophoresis-electrospray mass spectrometry (CE/ESI-MS) has provided new insight into the characterization of mannooligosaccharide caps from lipoarabinomannans (LAMs), which are key molecules in the immunopathogenesis of tuberculosis. This analytical approach requires oligosaccharide labeling with the fluorophore 1-aminopyrene-3,6,8-trisulfonate (APTS) by reductive amination at the reducing ... More
Comprehensive assessment of N-glycans derived from a murine monoclonal antibody: a case for multimethodological approach.
AuthorsMechref Y, Muzikar J, Novotny MV
JournalElectrophoresis
PubMed ID15841499
'Highly efficient separation techniques, laser-induced fluorescence (LIF) detection, and different mass-spectrometric (MS) measurements were combined in a multimethodological scheme to perform a comprehensive structural characterization of N-linked oligosaccharides in a murine monoclonal antibody (immunoglobulin G (IgG(kappa))). Monosaccharide compositional analysis was carried out through a capillary electrophoresis (CE)-LIF method, in which ... More
Ultrafast analysis of oligosaccharides on microchip with light-emitting diode confocal fluorescence detection.
AuthorsDang F, Zhang L, Hagiwara H, Mishina Y, Baba Y
JournalElectrophoresis
PubMed ID12601743
'We have developed a new method for the high-speed separation and high-sensitivity detection of complex oligosaccharides based on microchip electrophoresis (nu-CE) with light-emitting diode (LED) confocal fluorescence detection. Oligosaccharides labeled with 8-aminopyrene-1,3,6-trisulfonate (APTS) were found to strongly adsorb to the surface of polymethylmethacrylate (PMMA) microchips. Accordingly, three classes of major ... More
Analysis of monosaccharide composition by capillary electrophoresis.
AuthorsGuttman A
JournalJ Chromatogr A
PubMed ID9129325
'The monosaccharide composition analysis described in this paper employs capillary electrophoretic separation of sugar monomers liberated from glycoproteins or oligosaccharides, by high temperature acidic hydrolysis. Trifluoroacetic acid was used for sialo- and neutral-sugar hydrolysis, and hydrochloric acid was used for amino-sugar hydrolysis. The neutral- and amino-sugars in the hydrolyzates were ... More
High-resolution carbohydrate profiling by capillary gel electrophoresis.
AuthorsGuttman A
JournalNature
PubMed ID8602248
Electrophoretic methods for the analysis of N-linked oligosaccharides.
AuthorsRaju TS
JournalAnal Biochem
PubMed ID10906231
High-sensitivity and high-resolution methods for glycoprotein analysis.
AuthorsAnumula KR
JournalAnal Biochem
PubMed ID10929803
Metabolic cytometry: monitoring oligosaccharide biosynthesis in single cells by capillary electrophoresis.
AuthorsKrylov SN, Arriaga EA, Chan NW, Dovichi NJ, Palcic MM
JournalAnal Biochem
PubMed ID10906232
Recent advances in capillary electrophoresis of carbohydrates.
AuthorsEl Rassi Z, Mechref Y
JournalElectrophoresis
PubMed ID8900934
Analysis of mono- and oligosaccharide isomers derivatized with 9-aminopyrene-1,4,6-trisulfonate by capillary electrophoresis with laser-induced fluorescence.
AuthorsChen FT, Evangelista RA
JournalAnal Biochem
PubMed ID7503418
A method of analysis of isomers of mono- and oligosaccharides derivatized with 9-aminopyrene-1,4,6-trisulfonate (APTS) by capillary electrophoresis (CE) in simple buffer systems is presented. The derivatization chemistry is performed at 75 degrees C for 1 h for most mono- and oligosaccharides. Sialyllactose was derivatized at 37 degrees C for 15 ... More
Acid-catalyzed reductive amination of aldoses with 8-aminopyrene-1,3,6-trisulfonate.
AuthorsEvangelista RA, Guttman A, Chen FT
JournalElectrophoresis
PubMed ID8900941
The reductive amination of monosaccharides with 8-aminopyrene-1,3,6-trisulfonate (APTS) in seven different organic acids including the commonly used acetic acid was investigated by capillary electrophoresis (CE) with laser-induced fluorescence (LIF) detection. The correlation between the yields of the saccharide-APTS adducts and pKa of the organic acid catalyst is consistent with general ... More
Glycome mapping on DNA sequencing equipment.
AuthorsLaroy W, Contreras R, Callewaert N,
JournalNat Protoc
PubMed ID17406262
Here we provide a detailed protocol for the analysis of protein-linked glycans on DNA sequencing equipment. This protocol satisfies the glyco-analytical needs of many projects and can form the basis of 'glycomics' studies, in which robustness, high throughput, high sensitivity and reliable quantification are of paramount importance. The protocol routinely ... More
Analysis of 1-aminopyrene-3,6,8-trisulfonate-derivatized oligosaccharides by capillary electrophoresis with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.
AuthorsSuzuki H, Müller O, Guttman A, Karger BL
JournalAnal Chem
PubMed ID9375516
Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), has been used for structure characterization of 1-aminopyrene-3,6,8-trisulfonate (APTS)-derivatized oligosaccharides previously separated by capillary electrophoresis (CE). The resolved components were first isolated by employing an automated high-resolution fraction collector. Using on-probe sample cleanup with a cation-exchange resin and a matrix mixture of ... More
Mannosylated lipoarabinomannans inhibit IL-12 production by human dendritic cells: evidence for a negative signal delivered through the mannose receptor.
AuthorsNigou J, Zelle-Rieser C, Gilleron M, Thurnher M, Puzo G
JournalJ Immunol
PubMed ID11390501
IL-12 is a key cytokine in directing the development of type 1 Th cells, which are critical to eradicate intracellular pathogens such as Mycobacterium tuberculosis. Here, we report that mannose-capped lipoarabinomannans (ManLAMs) from Mycobacterium bovis bacillus Calmette-Guérin and Mycobacterium tuberculosis inhibited, in a dose-dependent manner, the LPS-induced IL-12 production by ... More
Comparison of fluorescence energy transfer primers with different donor-acceptor dye combinations.
AuthorsHung SC, Mathies RA, Glazer AN
JournalAnal Biochem
PubMed ID9448839
Fluorescence energy transfer (ET) primers are far superior to single dye-labeled primers as labels for DNA sequencing and polymerase chain reaction amplification. We compare here ET primers with different donor and acceptor dye combinations with respect to the relative acceptor fluorescence emission intensity and the amount of residual donor fluorescence ... More
Carbohydrate chromatography: towards yoctomole sensitivity.
AuthorsDavies MJ, Hounsell EF
JournalBiomed Chromatogr
PubMed ID8949909
In attempting to analyse complex carbohydrates by HPLC or capillary electrophoresis (CE) the chromatographer is faced with many challenges. The diversity of oligosaccharide structures described the date and their poor resolution on many chromatographic systems present a significant challenge to the chromatographer. In addition to this, the non-chromogenic nature of ... More
Capillary gel electrophoresis separation of high-mannose type oligosaccharides derivatized by 1-aminopyrene-3,6,8-trisulfonic acid.
AuthorsGuttman A, Pritchett T
JournalElectrophoresis
PubMed ID8586063
In this paper we report the capillary gel electrophoresis separation of 1-aminopyrene-3,6,8-trisulfonic acid (APTS) labeled oligosaccharides, released enzymatically from bovine pancreatic ribonuclease B. The released and labeled high-mannose structures were identified by spiking the separated peaks with the appropriate commercially available individual oligosaccharides. Baseline separation of the three positional isomers ... More
High-resolution capillary gel electrophoresis of reducing oligosaccharides labeled with 1-aminopyrene-3,6,8-trisulfonate.
AuthorsGuttman A, Chen FT, Evangelista RA, Cooke N
JournalAnal Biochem
PubMed ID8789724
High-resolution capillary gel electrophoresis was used for the separation of oligosaccharides labeled with a novel fluorophore 1-aminopyrene-3,6,8-trisulfonate (APTS) at the reducing termini by reductive amination. The APTS-saccharide adducts were detected by laser-induced fluorescence with excitation by the 488-nm Ar-ion laser and a 520-nm emission filter. The stoichiometry of labeling is ... More
Separation of 1-aminopyrene-3,6,8-trisulfonate-labeled asparagine-linked fetuin glycans by capillary gel electrophoresis.
AuthorsGuttman A, Chen FT, Evangelista RA
JournalElectrophoresis
PubMed ID8900952
Asparagine-linked glycans of bovine fetuin were separated by capillary gel electrophoresis after enzymatic release (peptide-N-glycosidase F) and labeling via reductive amination by a fluorescent dye, 1-aminopyrene-3,6-8-trisulfonate (APTS). At low separation pH (2.5) only two dominant peaks were observed. Increasing the separation buffer pH to 4.75 resulted in complete separation of ... More
A novel lipoarabinomannan from the equine pathogen Rhodococcus equi. Structure and effect on macrophage cytokine production.
Rhodococcus equi is a major cause of foal morbidity and mortality. We have investigated the presence of lipoglycan in this organism as closely related bacteria, notably Mycobacterium tuberculosis, produce lipoarabinomannans (LAM) that may play multiple roles as virulence determinants. The lipoglycan was structurally characterized by gas chromatography-mass spectrometry following permethylation, ... More
High resolution slab gel electrophoresis of 8-amino-1,3, 6-pyrenetrisulfonic acid (APTS) tagged oligosaccharides using a DNA sequencer.
AuthorsO'Shea MG, Morell MK
JournalElectrophoresis
PubMed ID8738327
A novel electrophoretic method for the analysis of oligosaccharides using DNA sequencer technology is illustrated using malto-oligosaccharide distributions obtained following isoamylase digestion of glycogen, wheat starch and potato starch. The debranched starches were derivatized at the reducing and with the charged fluorophore 8-amino-1,3,6-pyrenetrisulfonic acid (APTS). This highly reproducible method provides ... More
Screening method of carbohydrate-binding proteins in biological sources by capillary affinity electrophoresis and its application to determination of Tulipa gesneriana agglutinin in tulip bulbs.
AuthorsNakajima K, Kinoshita M, Oda Y, Masuko T, Kaku H, Shibuya N, Kakehi K
JournalGlycobiology
PubMed ID15148295
We developed capillary affinity electrophoresis (CAE) to analyze the molecular interaction between carbohydrate chains and proteins in solution state. A mixture of oligosaccharides derived from a glycoprotein was labeled with 8-aminopyrene-1,3,6-trisulfonate (APTS), and used as glycan library without isolation. Interaction of a carbohydrate-binding protein with each oligosaccharide in the mixture ... More
Determination of glycosaminoglycan monosaccharides by capillary electrophoresis using laser-induced fluorescence detection.
AuthorsRuiz-Calero V, Puignou L, Galceran MT
JournalJ Chromatogr B Analyt Technol Biomed Life Sci
PubMed ID12798179
A newly developed capillary electrophoretic method using laser-induced fluorescence detection (CE-LIF) for the analysis of monosaccharides released from acid hydrolysis of glycosaminoglycans was studied. The method was compared with a previously published method using indirect LIF detection (CE-ILIF). For the CE-LIF method, electrophoretic conditions for the separation of the monosaccharides ... More
The molecular deposition of transgenically modified starch in the starch granule as imaged by functional microscopy.
AuthorsBlennow A, Hansen M, Schulz A, Jørgensen K, Donald AM, Sanderson J
JournalJ Struct Biol
PubMed ID14572478
The molecular deposition of starch extracted from normal plants and transgenically modified potato lines was investigated using a combination of light microscopy, environmental scanning electron microscopy (ESEM) and confocal laser scanning microscopy (CLSM). ESEM permitted the detailed (10 nm) topographical analysis of starch granules in their hydrated state. CLSM could ... More
Capillary electrophoresis analysis of glucooligosaccharide regioisomers.
AuthorsJoucla G, Brando T, Remaud-Simeon M, Monsan P, Puzo G
JournalElectrophoresis
PubMed ID15004847
Complex gluco-oligosaccharide mixtures of two regioisomer series were successfully separated by CE. The gluco-oligosaccharide series were synthesized, employing a dextransucrase from Leuconostoc mesenteroides NRRL B-512F, by successive glucopyranosyl transfers from sucrose to the acceptor glucose or maltose. The glucosyl transfer to both acceptors, occurring through the formation of alpha1-->6 linkages, ... More
Structural characterization of Mycobacterium tuberculosis lipoarabinomannans by the combination of capillary electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.
AuthorsLudwiczak P, Brando T, Monsarrat B, Puzo G
JournalAnal Chem
PubMed ID11393859
Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) associated with capillary electrophoresis (CE) has been used for structural characterization of mannooligosaccharide caps from Mycobacterium tuberculosis H37rv mannosylated lipoarabinomannans (ManLAMs). The mannooligosaccharide caps were released by mild acid hydrolysis, labeled with 1-aminopyrene-3,6,8-trisulfonate (APTS) prior to being separated by CE, collected, and analyzed ... More
The phosphatidyl-myo-inositol anchor of the lipoarabinomannans from Mycobacterium bovis bacillus Calmette Guérin. Heterogeneity, structure, and role in the regulation of cytokine secretion.
AuthorsNigou J, Gilleron M, Cahuzac B, Bounéry JD, Herold M, Thurnher M, Puzo G
JournalJ Biol Chem
PubMed ID9287310
Lipoarabinomannans are major mycobacterial antigens capable of modulating the host immune response; however, the molecular basis underlying the diversity of their immunological properties remain an open question. In this study a new extraction and purification approach was successfully applied to isolate ManLAMs (lipoarabinomannans with mannosyl extensions) from bacillus Calmette Guérin ... More
Quantitative analysis of sugar constituents of glycoproteins by capillary electrophoresis.
AuthorsChen FT, Dobashi TS, Evangelista RA
JournalGlycobiology
PubMed ID9751791
A method for quantitative analysis of monosaccharides including N-acetylneuraminic acid derived from sialic acid-containing oligosaccharides and glycoproteins is presented. The analysis is based on the combination of chemical and enzymatic methods coupled with capillary electrophoretic (CE) separation and laser-induced fluorescence (LIF) detection. The present method utilizes a simplified acid hydrolysis ... More