Application of 2- Aminopyridine Fluorescence Labeling to Glycosaminoglycans1

Kon, Atsushi; Takagaki, Keiichi; Kawasaki, Hitoshi; Nakamura, Toshiya; Endo, Masahiko

doi:10.1093/oxfordjournals.jbchem.a123531

Cited by 67 publications

(37 citation statements)

References 2 publications

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“…The pyridylamination of the reducing termini of sugar chains has been useful for structural analysis and metabolic studies of N-or Oglycosidically linked sugar chains (18). We used a labeling system of DSS in order to investigate the metabolism of DSS.…”

Section: Methodsmentioning

confidence: 99%

Proteus mirabilis sp. intestinal microflora grow in a dextran sulfate sodium-rich environment

Araki¹

2009

Int J Mol Med

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Section: Methodsmentioning

confidence: 99%

Proteus mirabilis sp. intestinal microflora grow in a dextran sulfate sodium-rich environment

Araki¹

2009

Int J Mol Med

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“…Pyridylamination of Oligosaccharides-Fluorescence labeling with PA of the reducing terminal sugar of oligosaccharides was performed as described previously (20), based on the method of Hase et al (21).…”

Section: Methodsmentioning

confidence: 99%

Domain Structure of Chondroitin Sulfate E Octasaccharides Binding to Type V Collagen

Takagaki

Munakata

Kakizaki

et al. 2002

Journal of Biological Chemistry

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We demonstrated previously that chondroitin sulfate E (ChS-E) binds to type V collagen (Munakata, H., Takagaki, K., Majima, M., and Endo, M. (1999) Glycobiology 9, 1023-1027). In this study, we investigated the structure and binding of ChS-E oligosaccharides. Eleven oligosaccharides were isolated from ChS-E by gel filtration chromatography and anion-exchange high performance liquid chromatography after hydrolysis with testicular hyaluronidase. Separately, seven oligosaccharides were custom synthesized using the transglycosylation reaction of testicular hyaluronidase. Structural analysis was performed by enzymatic digestions in conjunction with high performance liquid chromatography and mass spectrometry. This library of 18 oligosaccharides was used as a source of model molecules to clarify the structural requirements for binding to type V collagen. Binding was analyzed by a biosensor based on surface plasmon resonance. The results indicated that to bind to type V collagen the oligosaccharides must have the following carbohydrate structures: 1) octasaccharide or larger in size; 2) a continuous sequence of three GlcA␤1-3GalNAc(4S,6S) units; 3) a GlcA␤1-3GalNAc(4S,6S) unit, GlcA␤1-3GalNAc(4S) unit or GlcA␤1-3GalNAc(6S) unit at the reducing terminal; 4) a GlcA␤1-3GalNAc(4S,6S) unit at the nonreducing terminal. It is likely that these characteristic oligosaccharide sequences play key roles in cell adhesion and extracellular matrix assembly.Proteoglycan exists in tissues of many mammalian species and is widely distributed in the cell surface and extracellular matrix. It is known that its carbohydrate chain, chondroitin sulfate (ChS), 1 has various important biological activities in areas such as cell migration, recognition, and morphogenesis (1-5). Recently, greater attention has been directed toward the functions of ChS in the brain, optic nerves, and chondrocytes (6 -8). ChS has many types of structural domains that are known to participate in specific physiological functions. However, the relationship between the biological function and structure of ChS domains is not yet fully understood. Recently, we used a surface plasmon resonance (SPR) biosensor to investigate the interaction of glycosaminoglycans (GAGs) with collagens and glycoproteins from the extracellular matrix (9). It was found that chondroitin sulfate E (ChS-E) has specific affinity for type V collagen, and that a novel characteristic sequence included in ChS-E is probably involved in binding to type V collagen. ChS-E from squid cartilage consists mainly of the disaccharide units GlcA␤1-3GalNAc(4S,6S), GlcA␤1-3GalNAc(4S), GlcA␤1-3GalNAc(6S), and GlcA␤1-3GalNAc, where 4S and 6S represent 4-O-and 6-O-sulfate, respectively (10, 11). Thus, since ChS-E has a variety of structures, it is likely that some specific feature of its structure is necessary for binding to type V collagen. However, the structural features within ChS-E that bind to type V collagen remain unclear.In the present work, we have prepared various oligosaccharides by testicular hyaluronidase...

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“…Fluorolabeling of the reducing ends of saturated ⁄ unsaturated HA oligosaccharides with PA was performed by a modification of the method of Hase et al [20], as described in our previous report [21]. In order to eliminate contamination of the substrates used with non-PA oligosaccharides, PA oligosaccharides were purified by HPLC, using a polyamine II column.…”

Section: Pyridylamination Of Ha Oligosaccharidesmentioning

confidence: 99%

Mechanism for the hydrolysis of hyaluronan oligosaccharides by bovine testicular hyaluronidase

Kakizaki¹,

Ibori²,

Kojima

et al. 2010

The FEBS Journal

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Synthetic hyaluronan oligosaccharides with defined structures and their pyridylaminated derivatives were used to investigate the mechanism of hydrolysis of hyaluronan by bovine testicular hyaluronidase. The products of the hydrolysis were analyzed by HPLC and ion‐spray mass spectroscopy (MS). It was confirmed that the minimum substrate for bovine testicular hyaluronidase is the hyaluronan hexasaccharide, even though it is a poor substrate that is barely cleaved, even on prolonged incubation. When hyaluronan octasaccharide was the substrate, increasing amounts of tetrasaccharide and hexasaccharide were produced with increasing time of incubation. Whereas disaccharide was not detectable in the reaction mixture by HPLC, MS analysis revealed trace amounts. The data suggest that the enzyme generates a disaccharide intermediate from hyaluronan oligosaccharide, the majority of which is transferred to the nonreducing ends of other oligosaccharides, only traces being released as free disaccharide. When hyaluronan octasaccharide, with an unsaturated glucuronic acid at the nonreducing end, was used as a substrate, only a tetrasaccharide was detected by HPLC. However, MS showed that the product was a mixture of equal amounts of two tetrasaccharides, one with and the other without the unsaturated glucuronic acid. This suggests that, in the case of substrates with a double bond at the nonreducing end, a tetrasaccharide is cleaved off instead of a disaccharide. The results of the experiments with pyridylaminated oligosaccharides were entirely consistent with these conclusions, and in addition showed the importance of the reducing end of the substrate for the enzyme to recognize the length of the saccharide.

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Application of 2- Aminopyridine Fluorescence Labeling to Glycosaminoglycans1

Cited by 67 publications

References 2 publications

Proteus mirabilis sp. intestinal microflora grow in a dextran sulfate sodium-rich environment

Proteus mirabilis sp. intestinal microflora grow in a dextran sulfate sodium-rich environment

Domain Structure of Chondroitin Sulfate E Octasaccharides Binding to Type V Collagen

Mechanism for the hydrolysis of hyaluronan oligosaccharides by bovine testicular hyaluronidase

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