The use of polyacrylamide gel electrophoresis for the analysis of acidic glycans labeled with the fluorophore 2-aminoacridone.
AuthorsJackson P, Pluskal MG, Skea W
JournalElectrophoresis
PubMed ID7813394
'A method is described for the electrophoretic analysis of fluorophore-labeled acidic glycans. Various glycoproteins were treated with the enzyme, peptide N-glycosidase F, to release mixtures of asparagine-linked glycans that were then labeled with the fluorophore 2-aminoacridone, and the resulting derivatives electrophoresed in high-density polyacrylamide gels. The acidic glycans were separated ... More
Microanalysis of glycosaminoglycan-derived disaccharides labeled with the fluorophore 2-aminoacridone by capillary electrophoresis and high-performance liquid chromatography.
AuthorsKitagawa H, Kinoshita A, Sugahara K
JournalAnal Biochem
PubMed ID8600818
'A series of disaccharides derived from chondroitin sulfate and heparin/heparan sulfate were derivatized at their reducing ends with the fluorophore 2-aminoacridone. The resulting labeled compounds derived from chondroitin sulfate or heparin/heparan sulfate were well separated and could be quantified by capillary electrophoresis and HPLC with a fluorescence detector. This method ... More
Electrospray mass spectrometry and fragmentation of N-linked carbohydrates derivatized at the reducing terminus.
AuthorsHarvey DJ
JournalJ Am Soc Mass Spectrom
PubMed ID11014452
'Derivatives were prepared from N-linked glycans by reductive amination from 2-aminobenzamide, 2-aminopyridine, 3-aminoquinoline, 2-aminoacridone, 4-amino-N-(2-diethylaminoethyl)benzamide, and the methyl, ethyl, and butyl esters of 4-aminobenzoic acid. Their electrospray and collision-induced dissociation (CID) fragmentation spectra were examined with a Q-TOF mass spectrometer. The strongest signals were obtained from the [M + Na]+ ... More
ADAMTS4 (aggrecanase-1) activation on the cell surface involves C-terminal cleavage by GPI-anchored MT4-MMP and binding of the activated proteinase to chondroitin sulfate and heparan sulfate on syndecan-1.
'C-terminal truncation of ADAMTS-4 from the p68 to the p53 form is required for activation of its capacity to cleave the Glu373-Ala374 interglobular domain bond of aggrecan. In transfected human chondrosarcoma cells this process is not autoproteolytic since the same products form with an inactive mutant of ADAMTS4 and truncation ... More
Minimizing cationization effects in the analysis of complex mixtures of oligosaccharides.
AuthorsNorth S, Okafo G, Birrell H, Haskins N, Camilleri P
JournalRapid Commun Mass Spectrom
PubMed ID9364792
'We outline simple methodology for the rapid and selective analysis of 2-aminoacridone (2-AMAC) derivatised oligosaccharides by matrix-assisted laser desorption/ionization mass spectrometry. This involves the addition of small amounts of lithium chloride to the matrix before evaporation of solvent and crystallization. Signals mainly attributed to proton, sodium and potassium adducts are ... More
Large-scale preparation, purification, and characterization of hyaluronan oligosaccharides from 4-mers to 52-mers.
AuthorsTawada A, Masa T, Oonuki Y, Watanabe A, Matsuzaki Y, Asari A
JournalGlycobiology
PubMed ID12122023
'Hyaluronan (HA) was depolymerized by partial digestion with testicular hyaluronidase and separated into size-uniform HA oligosaccharides from 4-mers to 52-mers by anion exchange chromatography after removal of the hyaluronidase. The purity and size of each HA oligosaccharide was confirmed by using HPLC analyses, FACE, and ESI-MS. (1)H and (13)C NMR ... More
Microanalysis of enzyme digests of hyaluronan and chondroitin/dermatan sulfate by fluorophore-assisted carbohydrate electrophoresis (FACE).
AuthorsCalabro A, Benavides M, Tammi M, Hascall VC, Midura RJ
JournalGlycobiology
PubMed ID10704526
'Hyaluronan and chondroitin/dermatan sulfate are glycosaminoglycans that play major roles in the biomechanical properties of a wide variety of tissues, including cartilage. A chondroitin/dermatan sulfate chain can be divided into three regions: (1) a single linkage region oligosaccharide, through which the chain is attached to its proteoglycan core protein, (2) ... More
A detailed analysis of neutral and acidic carbohydrates in human milk.
AuthorsCharlwood J, Tolson D, Dwek M, Camilleri P
JournalAnal Biochem
PubMed ID10469497
'Reverse- and normal-phase chromatography have been used to separate a number of standard human milk oligosaccharides derivatized via a reductive amination reaction with 2-aminoacridone (2-AMAC). Analytes were detected by spectrofluorimetry and injected simultaneously with a hydrolyzed dextran ladder derivatized with methyl-4-aminobenzoate. The latter probe does not fluoresce at the wavelength ... More
Identification and biosynthesis of cyclic enterobacterial common antigen in Escherichia coli.
AuthorsErbel PJ, Barr K, Gao N, Gerwig GJ, Rick PD, Gardner KH
JournalJ Bacteriol
PubMed ID12618464
'Phosphoglyceride-linked enterobacterial common antigen (ECA(PG)) is a cell surface glycolipid that is synthesized by all gram-negative enteric bacteria. The carbohydrate portion of ECA(PG) consists of linear heteropolysaccharide chains comprised of the trisaccharide repeat unit Fuc4NAc-ManNAcA-GlcNAc, where Fuc4NAc is 4-acetamido-4,6-dideoxy-D-galactose, ManNAcA is N-acetyl-D-mannosaminuronic acid, and GlcNAc is N-acetyl-D-glucosamine. The potential reducing ... More
Differences in the acid-labile component of Candida albicans mannan from hydrophobic and hydrophilic yeast cells.
AuthorsMasuoka J, Hazen KC
JournalGlycobiology
PubMed ID10536044
'Cell surface hydrophobicity of the opportunistic fungal pathogen Candida albicans has been linked to the level of cell wall protein glycosylation. Previous work demonstrated that outer chain mannosylation, rather than overall glycosylation, correlated with cell surface hydrophobicity. These studies further suggested that the phosphodiester-linked, acid-labile beta-1,2-mannan was the correlating element. ... More
Microscale analysis of mucin-type O-glycans by a coordinated fluorophore-assisted carbohydrate electrophoresis and mass spectrometry approach.
AuthorsRobbe C, Capon C, Flahaut C, Michalski JC
JournalElectrophoresis
PubMed ID12601728
'The total glycan moiety was released in a single step from native glycoproteins by a nonreductive beta-elimination procedure. The generated oligosaccharides were further derivatized either with the hydrophobic fluorophore 2-aminoacridone (AMAC) or the charged 8-aminonaphthalene-1,3,6-trisulfonic acid (ANTS) fluorophore, and the resulting fluorescent derivatives were separated according to their hydrodynamic size ... More
The analysis of fluorophore-labeled glycans by high-resolution polyacrylamide gel electrophoresis.
AuthorsJackson P
JournalAnal Biochem
PubMed ID8179179
'Many important biological processes are mediated by the carbohydrate moieties of glycoconjugates. Their structural determination presents considerable difficulties because of both their complex structures and the small quantities which are available from biological sources. In order to address some of the problems in this field a new method of glycan ... More
Epidermal growth factor activates hyaluronan synthase 2 in epidermal keratinocytes and increases pericellular and intracellular hyaluronan.
'Hyaluronan is an abundant and rapidly turned over matrix molecule between the vital cell layers of the epidermis. In this study, epidermal growth factor (EGF) induced a coat of hyaluronan and a 3-5-fold increase in its rate of synthesis in a rat epidermal keratinocyte cell line that has retained its ... More
Microanalysis of glycosaminoglycan-derived oligosaccharides labeled with a fluorophore 2-aminobenzamide by high-performance liquid chromatography: application to disaccharide composition analysis and exosequencing of oligosaccharides.
AuthorsKinoshita A, Sugahara K
JournalAnal Biochem
PubMed ID10222012
'A series of disaccharides derived from chondroitin sulfate and heparin/heparan sulfate were derivatized at their reducing ends with a fluorophore 2-aminobenzamide to develop a sensitive microanalytical method for glycosaminoglycans. The resulting labeled compounds derived from chondroitin sulfate or heparin/heparan sulfate were well-separated and quantified by HPLC equipped with a fluorescence ... 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
Analysis of glycosphingolipids by fluorophore-assisted carbohydrate electrophoresis using ceramide glycanase from Mercenaria mercenaria.
AuthorsBasu SS, Dastgheib-Hosseini S, Hoover G, Li Z, Basu S
JournalAnal Biochem
PubMed ID7856860
'Polyacrylamide gel electrophoresis of fluorophore-labeled saccharides is a simple and sensitive separation method for glycan analysis. This method requires an aldehydic reducing carbon on the saccharide in order to react with the amine group of the fluorophore (8-aminonaphthalene-1,3,6-trisulfonic acid or 2-aminoacridone). We have used exoglycosidase-free ceramide glycanase from hard-shelled clam ... More
Ultrasensitive capillary electrophoresis of sulfated disaccharides in chondroitin/dermatan sulfates by laser-induced fluorescence after derivatization with 2-aminoacridone.
AuthorsLamari F, Theocharis A, Hjerpe A, Karamanos NK
JournalJ Chromatogr B Biomed Sci Appl
PubMed ID10437680
'An ultrasensitive capillary electrophoretic method for separating the variously sulfated chondroitin/dermatan sulfate-derived delta-disaccharides after digestion with chondro/dermatolyases and derivatization with the fluorophore 2-aminoacridone is described. All known mono-, di- and tri-sulfated delta-disaccharides were completely separated using 15 mM orthophosphate buffer (pH 3.0) at 20 kV without any interference of the ... More
The glycosylation of IgA produced by murine B cells is altered by Th2 cytokines.
AuthorsChintalacharuvu SR, Emancipator SN
JournalJ Immunol
PubMed ID9278322
'We sought to determine whether selected cytokines, known to profoundly increase proliferation and/or production of Ig by B cells and their progeny, also have as yet unrecognized effects upon IgA glycosylation. For these studies, we selected CH12LX mouse B lymphoma cells, a widely used model of B cell differentiation. Glycosylation ... More
Analysis of neutral saccharides in human milk derivatized with 2-aminoacridone by capillary electrophoresis with laser-induced fluorescence detection.
AuthorsSong JF, Weng MQ, Wu SM, Xia QC
JournalAnal Biochem
PubMed ID11969197
Efficient carbohydrate release, purification, and derivatization.
AuthorsCharlwood J, Birrell H, Camilleri P
JournalAnal Biochem
PubMed ID9750134
High-resolution polyacrylamide gel electrophoresis of fluorophore-labeled reducing saccharides.
AuthorsJackson P
JournalMethods Enzymol
PubMed ID8139500
High-sensitivity and high-resolution methods for glycoprotein analysis.
AuthorsAnumula KR
JournalAnal Biochem
PubMed ID10929803
Recent advances in capillary electrophoresis of carbohydrates.
AuthorsEl Rassi Z, Mechref Y
JournalElectrophoresis
PubMed ID8900934
Direct structural analysis of 2-aminoacridone derivatized oligosaccharides by high-performance liquid chromatography/mass spectrometric detection.
AuthorsCamilleri P, Tolson D, Birrell H
JournalRapid Commun Mass Spectrom
PubMed ID9474802
In this preliminary study linear and branched oligosaccharides, derivatized with 2-aminoacridone, have been separated by reverse-phase high-performance liquid chromatography, and detected by positive-ion electrospray mass spectrometry. Tandem Mass spectra of the protonated carbohydrate analytes gave simple fragmentation patterns from which sequence information could be readily obtained. ... More
Fingerprinting of glycans as their 2-aminoacridone derivatives by capillary electrophoresis and laser-induced fluorescence.
Capillary electrophoresis and laser-induced fluorescence detection have been used to fingerprint the 2-aminoacridone derivatives of complex glycans released from bovine fetuin and human IgG monoclonal antibodies. The utility of this method in distinguishing between N- and O-linked oligosaccharides and in determining the presence of sialic acid residues in glycan mixtures ... More
Carbohydrate release from picomole quantities of glycoprotein and characterisation of glycans by high-performance liquid chromatography and mass spectrometry.
AuthorsCharlwood J, Birrell H, Camilleri P
JournalJ Chromatogr B Biomed Sci Appl
PubMed ID10574203
Samples of 5 to 20 microg of human IgG were subjected to dithiothreitol treatment to reduce disulphide bridges, followed by tryptic digestion. Glycans released from the tryptic peptide mixture by PNGase F digestion were then derivatised with 2-aminoacridone. Labelled oligosaccharides were separated by normal-phase high-performance liquid chromatography and individual components ... More
Profiling of 2-aminoacridone derivatised glycans by electrospray ionization mass spectrometry.
AuthorsCharlwood J, Langridge J, Tolson D, Birrell H, Camilleri P
JournalRapid Commun Mass Spectrom
PubMed ID9951411
Three different types of N-glycans, high-mannose, complex and hybrid, have been derivatised with 2-aminoacridone and analysed by electrospray mass spectrometry and tandem mass spectrometry (MS/MS) using a hybrid quadrupole orthogonal acceleration time-of-flight mass spectrometer, fitted with a nanoflow electrospray ion source. Relatively simple MS/MS fragmentation patterns have been observed for ... More
Two-dimensional chromatography in the analysis of complex glycans from transferrin.
AuthorsCharlwood J, Birrell H, Tolson D, Camilleri P
JournalAnal Chem
PubMed ID9666724
Oligosaccharides released from human transferrin have been derivatized with 2-aminoacridone (2-AMAC) prior to analysis by either reversed- or normal-phase high-performance liquid chromatography. These separation methods are complementary and allow distinction between isomeric mono- and disialylated oligosaccharides, which terminate with Sia alpha 2-6Gal or Sia alpha 2-3Gal at the nonreducing end. ... More
Simple differentiation between core-fucosylated and nonfucosylated glycans by capillary electrophoresis.
AuthorsOkafo GN, Burrow LM, Neville W, Truneh A, Smith RA, Reff M, Camilleri P
JournalAnal Biochem
PubMed ID8811881
Core-fucosylated glycans, derivatized with 2-aminoacridone, consistently migrate slower than the corresponding oligosaccharides which lack this fucose residue, using the micellar electrophoretic capillary chromatography conditions outlined in this study. alpha-Fucosidase digestion of glycans followed by CE analysis has allowed facile differentiation of these two classes of oligosaccharides and this methodology has ... More
A probe for the versatile analysis and characterization of N-linked oligosaccharides.
AuthorsCharlwood J, Birrell H, Gribble A, Burdes V, Tolson D, Camilleri P
JournalAnal Chem
PubMed ID10763240
A novel fluorescent probe, 3-(acetylamino)-6-aminoacridine (AA-Ac), has been synthesized and its applicability to the analysis of picomole levels of N-linked glycans investigated. AA-Ac was found to be an excellent derivatization reagent for N-linked glycans, giving at least twice the intensity of fluorescence as its predecessor 2-aminoacridone. AA-Ac-labeled glycans were analyzed ... More
Analysis of oligosaccharides by microbore high-performance liquid chromatography.
AuthorsCharlwood J, Birrell H, Bouvier ES, Langridge J, Camilleri P
JournalAnal Chem
PubMed ID10763242
A 1-mm microbore hydrophilic interaction column has been used for the separation of 2-aminoacridone (2-AMAC)-derivatized glycan mixtures, released from naturally occurring and recombinant proteins. Primary structure identification of the 2-AMAC glycan derivatives was carried out by HPLC using fluorescence and mass spectrometric detection. In some cases, enzymatic digestion of the ... More
Early indication of effects of puromycin aminonucleoside using a fluorimetric assay of 2-aminoacridone-derivatized carbohydrates in urine.
AuthorsCharlwood J, Birrell H, Tolson D, Connelly J, Camilleri P
JournalAnal Biochem
PubMed ID10906246
This paper describes a novel noninvasive method to study the changes in free carbohydrates excreted in urine as a result of toxicity in the rat induced by the administration of puromycin aminonucleoside (PAN). Urine samples were collected for 24 h prior to dosing and at 8, 24, and 32 h ... More
Structural characterisation of N-linked glycan mixtures by precursor ion scanning and tandem mass spectrometric analysis
AuthorsCharlwood J, Langridge J, Camilleri P
JournalRapid Commun Mass Spectrom
PubMed ID10407349
2-Aminoacridone (2-AMAC) labelled N-linked glycan pools were analysed directly by a hybrid quadrupole orthogonal acceleration time-of-flight mass spectrometer (Q-Tof) in the precursor ion scanning and tandem mass spectrometry (MS/MS) modes. The use of a precursor ion scanning strategy on this instrument provides a rapid and sensitive method of screening glycan ... More
A coordinated high-performance liquid chromatographic, capillary electrophoretic, and mass spectrometric approach for the analysis of oligosaccharide mixtures derivatized with 2-aminoacridone.
AuthorsOkafo G, Burrow L, Carr SA, Roberts GD, Johnson W, Camilleri P
JournalAnal Chem
PubMed ID8972625
Glycans derivatized with 2-aminoacridone have been analyzed consecutively by reverse-phase high-performance liquid chromatography (HPLC) and micellar electrokinetic capillary chromatography (MECC). The 2-aminoacridone derivatizing agent used in the present study is highly hydrophobic and is well separated from the glycan derivatives in both separation techniques, ensuring that excess reagent does not ... More
High-performance liquid chromatographic analysis of complex N-linked glycans derivatized with 2-aminoacridone.
AuthorsOkafo G, Langridge J, North S, Organ A, West A, Morris M, Camilleri P
JournalAnal Chem
PubMed ID9414612
2-Aminoacridone (2-AMAC) has been used to derivatize mixtures of N-linked oligosaccharides released from alpha(1)-acid glycoprotein and immunoglobulin G. In each case, the HPLC profile obtained for the derivatized glycans was compared to that obtained after digestion with sialidase and a two-enzyme array system made up of sialidase and alpha-fucosidase, prior ... More
Development of a functional bioassay for arylsulfatase B using the natural substrates of the enzyme.
AuthorsPungor E, Hague CM, Chen G, Lemontt JF, Dvorak-Ewell M, Prince WS,
JournalAnal Biochem
PubMed ID19682969
A functional bioassay has been developed for measuring the intracellular activity of recombinant human arylsulfatase B (rhASB) on its natural glycosaminoglycan (GAG) substrates, dermatan sulfate (DS), and chondroitin sulfate (CS) when the enzyme is taken up into cultured ASB-deficient human fibroblasts (GM00519). The enzyme ASB is a lysosomal exohydrolase, cleaving ... More
A chromatographic and mass spectrometric strategy for the analysis of oligosaccharides: determination of the glycan structures in porcine thyroglobulin.
AuthorsCharlwood J, Birrell H, Organ A, Camilleri P
JournalRapid Commun Mass Spectrom
PubMed ID10343413
Oligosaccharides released from porcine thyroglobulin were first derivatised with 2-aminoacridone (2-AMAC) and analysed by capillary electrophoresis to determine the complexity of this glycan pool. The same glycan mixture was then subjected to either a sialidase digest or a sialidase and fucosidase digest prior to derivatisation with 2-AMAC and analysis by ... More
High resolution and rapid analysis of branched oligosaccharides by capillary electrophoresis.
AuthorsCamilleri P, Harland GB, Okafo G
JournalAnal Biochem
PubMed ID8585606
The fluorophore 2-aminoacridone has been used to label a number of branched oligosaccharides previously released from various glycoproteins. Complex glycans were derivatized via their reducing end with this fluorophore by a Schiff's base mechanism followed by reduction to a secondary amine using sodium cyanoborohydride. This process of derivatization was carried ... More
Fluorometric continuous kinetic assay of alpha-chymotrypsin using new protease substrates possessing long-wave excitation and emission maxima.
AuthorsBaustert JH, Wolfbeis OS, Moser R, Koller E
JournalAnal Biochem
PubMed ID3407938
A direct and continuous kinetic method for the fluorometric determination of alpha-chymotrypsin and trypsin is described, and 2-aminoacridone (2-AA) is introduced as a promising new fluorophore in analytical biochemistry. N-Succinyl- and N-glutaryl-phenylalanine as well as N-benzoylarginine were coupled to 2-AA via a peptide bond and the resulting fluorogenic substrates are ... More
Extracellular glycosaminoglycans modify cellular trafficking of lipoplexes and polyplexes.
Authors Ruponen M; Rönkkö S; Honkakoski P; Pelkonen J; Tammi M; Urtti A;
JournalJ Biol Chem
PubMed ID11390375
It has been shown that extracellular glycosaminoglycans (GAGs) limit the gene transfer by cationic lipids and polymers. The purpose of this study was to clarify how interactions with anionic GAGs (hyaluronic acid and heparan sulfate) modify the cellular uptake and distribution of lipoplexes and polyplexes. Experiments on cellular DNA uptake ... More
Advancing monosaccharides as biomarkers: part I. Development of fluorophore-assisted carbohydrate-electrophoresis in Chironomus riparius.
AuthorsBentivegna CS
JournalAquat Toxicol
PubMed ID12297374
Fluorophore-assisted carbohydrate-electrophoresis (FACE) was developed as a bioassay for environmental stressors in larval Chironomus riparius. This quantitative technique involved acid hydrolysis and 2-aminoacridone labeling of monosaccharides followed by carbohydrate gel electrophoresis. Methods for carbohydrate isolation from whole tissue homogenates as well as migration distances for 23 different monosaccharides and 4 ... More
Polysaccharide analysis using carbohydrate gel electrophoresis: a method to study plant cell wall polysaccharides and polysaccharide hydrolases.
AuthorsGoubet F, Jackson P, Deery MJ, Dupree P
JournalAnal Biochem
PubMed ID11743692
A method to characterize plant cell wall polysaccharides is presented. The complexity of the polymer structures and the large number of different charged and uncharged monosaccharides that make up plant polysaccharides have previously made analysis technically demanding and laborious. Polysaccharide analysis using carbohydrate gel electrophoresis (PACE) relies on derivatization of ... More
Characterization of the glycosylation profiles of Alzheimer's beta -secretase protein Asp-2 expressed in a variety of cell lines.
AuthorsCharlwood J, Dingwall C, Matico R, Hussain I, Johanson K, Moore S, Powell DJ, Skehel JM, Ratcliffe S, Clarke B, Trill J, Sweitzer S, Camilleri P
JournalJ Biol Chem
PubMed ID11278492
Amyloid 39-42 beta -peptides are the main components of amyloid plaques found in the brain of Alzheimer's disease patients. Amyloid 39-42 beta-peptide is formed from amyloid precursor protein by the sequential action of beta- and gamma-secretases. Asp-2 is a transmembrane aspartic protease expressed in the brain, shown to have beta-secretase ... More
Polyacrylamide gel electrophoresis of reducing saccharides labeled with the fluorophore 2-aminoacridone: subpicomolar detection using an imaging system based on a cooled charge-coupled device.
AuthorsJackson P
JournalAnal Biochem
PubMed ID1776673
Numerous monosaccharides and oligosaccharides were derivatized at their reducing end groups with the fluorophore 2-aminoacridone. The resulting fluorescent compounds were separated by PAGE using two different buffer systems. One of these, a Tris borate buffer, enabled all of the fluorescent saccharide derivatives tested to be electrophoresed and various positional isomers, ... More
Liquid chromatographic assay for a glucose tetrasaccharide, a putative biomarker for the diagnosis of Pompe disease.
AuthorsAn Y, Young SP, Hillman SL, Van Hove JL, Chen YT, Millington DS
JournalAnal Biochem
PubMed ID11078593
A HPLC method associated with butyl-p-aminobenzoate derivatization has been developed for the analysis of a tetraglucose oligomer, Glcalpha1-6Glcalpha1-4Glcalpha1-4Glc, designated Glc(4), in biological fluids. This tetraglucose, normally excreted in the urine, has previously been shown to be elevated in a number of pathological conditions including Pompe disease (glycogen storage disease type ... More
Molecular cloning and characterization of Bifidobacterium bifidum 1,2-alpha-L-fucosidase (AfcA), a novel inverting glycosidase (glycoside hydrolase family 95).
AuthorsKatayama T, Sakuma A, Kimura T, Makimura Y, Hiratake J, Sakata K, Yamanoi T, Kumagai H, Yamamoto K
JournalJ Bacteriol
PubMed ID15262925
A genomic library of Bifidobacterium bifidum constructed in Escherichia coli was screened for the ability to hydrolyze the alpha-(1-->2) linkage of 2'-fucosyllactose, and a gene encoding 1,2-alpha-l-fucosidase (AfcA) was isolated. The afcA gene was found to comprise 1,959 amino acid residues with a predicted molecular mass of 205 kDa and ... More
Binding of a large chondroitin sulfate/dermatan sulfate proteoglycan, versican, to L-selectin, P-selectin, and CD44.
AuthorsKawashima H, Hirose M, Hirose J, Nagakubo D, Plaas AH, Miyasaka M
JournalJ Biol Chem
PubMed ID10950950
Here we show that a large chondroitin sulfate proteoglycan, versican, derived from a renal adenocarcinoma cell line ACHN, binds L-selectin, P-selectin, and CD44. The binding was mediated by the interaction of the chondroitin sulfate (CS) chain of versican with the carbohydrate-binding domain of L- and P-selectin and CD44. The binding ... More
Hoxb13 knockout adult skin exhibits high levels of hyaluronan and enhanced wound healing.
AuthorsMack JA, Abramson SR, Ben Y, Coffin JC, Rothrock JK, Maytin EV, Hascall VC, Largman C, Stelnicki EJ
JournalFASEB J
PubMed ID12759339
In contrast to adult cutaneous wound repair, early gestational fetal cutaneous wounds heal by a process of regeneration, resulting in little or no scarring. Previous studies indicate that down-regulation of HoxB13, a member of the highly conserved family of Hox transcription factors, occurs during fetal scarless wound healing. No down-regulation ... More
Microanalysis of hyaluronan oligosaccharides by polyacrylamide gel electrophoresis and its application to assay of hyaluronidase activity.
AuthorsIkegami-Kawai M, Takahashi T
JournalAnal Biochem
PubMed ID12470675
We established a rapid, sensitive polyacrylamide gel electrophoresis (PAGE) method for the analysis of hyaluronan (HA) oligosaccharides. Using mini-slab gels, but not large-slab gels so far reported, HA oligosaccharides of 5 to more than 50 repeating disaccharide units could be separated into discrete ladder-like bands in a short electrophoresis time ... More
Keratinocyte growth factor stimulates migration and hyaluronan synthesis in the epidermis by activation of keratinocyte hyaluronan synthases 2 and 3.
AuthorsKarvinen S, Pasonen-Seppänen S, Hyttinen JM, Pienimäki JP, Törrönen K, Jokela TA, Tammi MI, Tammi R
JournalJ Biol Chem
PubMed ID14506240
Keratinocyte growth factor (KGF) activates keratinocyte migration and stimulates wound healing. Hyaluronan, an extracellular matrix glycosaminoglycan that accumulates in wounded epidermis, is known to promote cell migration, suggesting that increased synthesis of hyaluronan might be associated with the KGF response in keratinocytes. Treatment of monolayer cultures of rat epidermal keratinocytes ... More
Rapid activation of glycogen phosphorylase by the endoplasmic reticulum unfolded protein response.
AuthorsGill A, Gao N, Lehrman MA
JournalJ Biol Chem
PubMed ID12223475
Endoplasmic reticulum (ER) stress is associated with misfolding of ER proteins and triggers the unfolded protein response (UPR). The UPR, in turn, helps restore normal ER function. Since fastidious N-linked glycosylation is critical for folding of most ER proteins, this study examined whether metabolic interconversions of precursors used for glycan ... More
Intracellular accumulation of secreted proteoglycans inhibits cationic lipid-mediated gene transfer. Co-transfer of glycosaminoglycans to the nucleus.
AuthorsBelting M, Petersson P
JournalJ Biol Chem
PubMed ID10383450
Molecules secreted by potential target cells may interfere with cationic lipid-mediated gene transfer. This has been studied using human lung fibroblasts and human epidermoid lung cancer cells. Secreted cell medium components caused a substantial decrease both in the uptake of cationic lipid-DNA complexes (2-4-fold) and in reporter gene expression (100-1000-fold). ... More
Identification and functions of chondroitin sulfate in the milieu of neural stem cells.
AuthorsIda M, Shuo T, Hirano K, Tokita Y, Nakanishi K, Matsui F, Aono S, Fujita H, Fujiwara Y, Kaji T, Oohira A
JournalJ Biol Chem
PubMed ID16373347
The behavior of cells is generally considered to be regulated by environmental factors, but the molecules in the milieu of neural stem cells have been little studied. We found by immunohistochemistry that chondroitin sulfate (CS) existed in the surroundings of nestin-positive cells or neural stem/progenitor cells in the rat ventricular ... More
Characterization of derivatization of sialic acid with 2-aminoacridone and determination of sialic acid content in glycoproteins by capillary electrophoresis and high performance liquid chromatography--ion trap mass spectrometry.
AuthorsChe FY, Shao XX, Wang KY, Xia QC
JournalElectrophoresis
PubMed ID10546830
A simple and highly sensitive capillary electrophoresis (CE) method for determining the content of N-acetylneuraminic acid (Neu5Ac) in glycoproteins was developed. Neu5Ac was derivatized with 2-aminoacridone (AMAC) by reductive amination, and the AMAC-Neu5Ac adduct could be readily separated from the other 11 AMAC-derivatized neutral and acidic monosaccharides usually present in ... More
Mononuclear leukocytes preferentially bind via CD44 to hyaluronan on human intestinal mucosal smooth muscle cells after virus infection or treatment with poly(I.C).
Authorsde La Motte CA, Hascall VC, Calabro A, Yen-Lieberman B, Strong SA
JournalJ Biol Chem
PubMed ID10521464
Pathological changes in inflammatory bowel disease include an increase in intestinal mucosal mononuclear leukocytes and hyperplasia of the muscularis mucosae smooth muscle cells (M-SMCs). Because virus infections have correlated with disease flare, we tested the response of cultured M-SMCs to respiratory syncytial virus, measles virus, and the viral analogue, poly(I.C). ... More
Changed lamellipodial extension, adhesion plaques and migration in epidermal keratinocytes containing constitutively expressed sense and antisense hyaluronan synthase 2 (Has2) genes.
AuthorsRilla K, Lammi MJ, Sironen R, Törrönen K, Luukkonen M, Hascall VC, Midura RJ, Hyttinen M, Pelkonen J, Tammi M, Tammi R
JournalJ Cell Sci
PubMed ID12186949
Hyaluronan is a major component of the epidermal extracellular matrix, is actively synthesized by keratinocytes and shows fast matrix turnover in the stratified epithelium. We probed the importance of hyaluronan synthesis in keratinocytes by establishing cell lines carrying the exogenous hyaluronan synthase 2 (Has2) gene in sense and antisense orientations ... More
Characterization of protein glycosylation using chip-based infusion nanoelectrospray linear ion trap tandem mass spectrometry.
AuthorsZhang S, Chelius D
JournalJ Biomol Tech
PubMed ID15190085
Mass spectrometry (MS) has the potential to revolutionize structural glycobiology and help in the understanding of how post-translation events such as glycosylation affect protein activities. Several approaches to determine the structure of glycopeptides have been used successfully including fast atom bombardment, matrix-assisted laser desorption ionization, and electrospray ionization with a ... More
Glycosaminoglycan synthesis and secretion by the retinal pigment epithelium: polarized delivery of hyaluronan from the apical surface.
AuthorsdeS Senanayake P Calabro A, Nishiyama K, Hu JG, Bok D, Hollyfield JG
JournalJ Cell Sci
PubMed ID11112703
Hyaluronan and chondroitin sulfate glycosaminoglycan secretion from retinal pigment epithelial cells was established in confluent cultures with high transepithelial resistance. Cell cultures were maintained on Millicell-PCF culture plates, which allow separation of culture medium exposed to apical and basal epithelial surfaces. Following various times in culture, apical and basal culture ... More
Keratocyte phenotype mediates proteoglycan structure: a role for fibroblasts in corneal fibrosis.
AuthorsFunderburgh JL, Mann MM, Funderburgh ML
JournalJ Biol Chem
PubMed ID12933807
In pathological corneas, accumulation of fibrotic extracellular matrix is characterized by proteoglycans with altered glycosaminoglycans that contribute to the reduced transparency of scarred tissue. During wound healing, keratocytes in the corneal stroma transdifferentiate into fibroblasts and myofibroblasts. In this study, molecular markers were developed to identify keratocyte, fibroblast, and myofibroblast ... More
Hyaluronan enters keratinocytes by a novel endocytic route for catabolism.
AuthorsTammi R, Rilla K, Pienimaki JP, MacCallum DK, Hogg M, Luukkonen M, Hascall VC, Tammi M
JournalJ Biol Chem
PubMed ID11451952
Hyaluronan synthesized in the epidermis has an exceptionally short half-life, indicative of its catabolism by epidermal keratinocytes. An intracellular pool of endogenously synthesized hyaluronan, from 1 to 20 fg/cell, inversely related to cell density, was observed in cultured rat epidermal keratinocytes. More than 80% of the intracellular hyaluronan was small ... More
Coupling of the dolichol-P-P-oligosaccharide pathway to translation by perturbation-sensitive regulation of the initiating enzyme, GlcNAc-1-P transferase.
AuthorsGao N, Lehrman MA
JournalJ Biol Chem
PubMed ID12176988
In mammalian cells, inhibition of translation interferes with synthesis of the lipid-linked oligosaccharide (LLO) Glc3Man9GlcNAc2-P-P-dolichol as measured with radioactive sugar precursors. Conflicting hypotheses have been proposed, and the fundamental basis for this regulation has remained elusive. Here, fluorophore-assisted carbohydrate electrophoresis (FACE) was used to measure LLO concentrations directly in cells ... More
Novel tetrasaccharides isolated from squid cartilage chondroitin sulfate E contain unusual sulfated disaccharide units GlcA(3-O-sulfate)beta1-3GalNAc(6-O-sulfate) or GlcA(3-O-sulfate)beta1-3GalNAc.
AuthorsKinoshita A, Yamada S, Haslam SM, Morris HR, Dell A, Sugahara K
JournalJ Biol Chem
PubMed ID9242620
We previously isolated novel tetrasaccharides containing 3-O-sulfated glucuronic acid from king crab cartilage chondroitin sulfate K and demonstrated that the disaccharide units containing 3-O-sulfated glucuronic acid were decomposed by chondroitinase ABC digestion (Sugahara, K., Tanaka, Y., Yamada, S., Seno, N., Kitagawa, H., Haslam, S. M., Morris, H. R., and Dell, ... More
Role of hyaluronan and reactive oxygen species in tissue kallikrein-mediated epidermal growth factor receptor activation in human airways.
AuthorsCasalino-Matsuda SM, Monzon ME, Conner GE, Salathe M, Forteza RM
JournalJ Biol Chem
PubMed ID14988406
In human airways, oxidative stress-induced submucosal gland cell hypertrophy and hyperplasia, histological features of chronic bronchitis, have been linked to epidermal growth factor receptor (EGFR) activation. To explore mechanisms of oxidative stress-induced EGFR activation and signaling, primary cultures of human tracheal submucosal gland (SMG) cells were used to assess EGFR ... More
Analyses of dolichol pyrophosphate-linked oligosaccharides in cell cultures and tissues by fluorophore-assisted carbohydrate electrophoresis.
AuthorsGao N, Lehrman MA
JournalGlycobiology
PubMed ID12070078
Lipid-linked oligosaccharides (LLOs) are the precursors of asparagine (N)-linked glycans, which are essential information carriers in many biological systems, and defects in LLO synthesis cause Type I congenital disorders of glycosylation. Due to the low abundance of LLOs and the limitations of the chemical and physical methods previously used to ... More
Novel methods for the preparation and characterization of hyaluronan oligosaccharides of defined length.
AuthorsMahoney DJ, Aplin RT, Calabro A, Hascall VC, Day AJ
JournalGlycobiology
PubMed ID11805075
Hyaluronan is a ubiquitous glycosaminoglycan of high molecular weight that acts as a structural component of extracellular matrices and mediates cell adhesion. There have been numerous recent reports that fragments of hyaluronan have different properties compared to the intact molecule. Though many of these results may be genuine, it is ... More
Influence of the secondary cell wall polymer on the reassembly, recrystallization, and stability properties of the S-layer protein from Bacillus stearothermophilus PV72/p2.
AuthorsSára M, Dekitsch C, Mayer HF, Egelseer EM, Sleytr UB
JournalJ Bacteriol
PubMed ID9696762
The high-molecular-weight secondary cell wall polymer (SCWP) from Bacillus stearothermophilus PV72/p2 is mainly composed of N-acetylglucosamine (GlcNAc) and N-acetylmannosamine (ManNAc) and is involved in anchoring the S-layer protein via its N-terminal region to the rigid cell wall layer. In addition to this binding function, the SCWP was found to inhibit ... More
Glycosaminoglycans and proteoglycans in normal mitral valve leaflets and chordae: association with regions of tensile and compressive loading.
AuthorsGrande-Allen KJ, Calabro A, Gupta V, Wight TN, Hascall VC, Vesely I
JournalGlycobiology
PubMed ID15044391
This study was designed to identify the specific proteoglycans and glycosaminoglycans (GAGs) in the leaflets and chordae of the mitral valve and to interpret their presence in relation to the tensile and compressive loads borne by these tissues. Leaflets and chordae from normal human mitral valves (n = 31, obtained ... More
Adaptation of FACE methodology for microanalysis of total hyaluronan and chondroitin sulfate composition from cartilage.
AuthorsCalabro A, Hascall VC, Midura RJ
JournalGlycobiology
PubMed ID10704527
Protocols for analyzing the fine structure of hyaluronan and chondroitin sulfate using fluorophore-assisted carbohydrate electrophoresis of 2-aminoacridone-derivatized hyaluronidase/chondroitinase digestion products were adapted for direct analysis of previously characterized cartilage-derived samples. The chondroitin sulfate disaccharide compositions for fetal and 68 year human aggrecan from FACE analyses were DeltaDi4S (50%), DeltaDi6S (43%), ... More
Hyaluronan structures synthesized by rat mesangial cells in response to hyperglycemia induce monocyte adhesion.
AuthorsWang A, Hascall VC
JournalJ Biol Chem
PubMed ID14679194
Mesangial expansion, the principal glomerular lesion in diabetic nephropathy, is preceded by a phenotypic activation and transient proliferation of the glomerular mesangial cells and by a prominent glomerular infiltration of monocytes and macrophages. Because this infiltration seems to play a key role in the subsequent mesangial matrix expansion, we tested ... More
Keratan sulfate disaccharide composition determined by FACE analysis of keratanase II and endo-beta-galactosidase digestion products.
AuthorsPlaas AH, West LA, Midura RJ
JournalGlycobiology
PubMed ID11588154
Many tissues contain glycoproteins and proteoglycans, which are substituted with N-or O-linked keratan sulfate, a glycosaminoglycan in which the lactosamine (-galbeta1,4glcNAc-) disaccharide backbone is variably modified by sulfation, fucosylation, and sialylation. We report here a rapid, sensitive, and quantitative procedure for obtaining a complete disaccharide compositional analyses for keratan sulfates ... More
The S-layer proteins of two Bacillus stearothermophilus wild-type strains are bound via their N-terminal region to a secondary cell wall polymer of identical chemical composition.
AuthorsEgelseer EM, Leitner K, Jarosch M, Hotzy C, Zayni S, Sleytr UB, Sára M
JournalJ Bacteriol
PubMed ID9515918
Two Bacillus stearothermophilus wild-type strains were investigated regarding a common recognition and binding mechanism between the S-layer protein and the underlying cell envelope layer. The S-layer protein from B. stearothermophilus PV72/p6 has a molecular weight of 130,000 and assembles into a hexagonally ordered lattice. The S-layer from B. stearothermophilus ATCC ... More