Chimeric Glycosaminoglycan Oligosaccharides Synthesized by Enzymatic Reconstruction and Their Use in Substrate Specificity Determination of Streptococcus Hyaluronidase
| Autor: | Masahiko Endo, Mitsuo Majima, Kelichi Takagaki, Hidekazu Munakata, lkuko Kakizaki |
|---|---|
| Rok vydání: | 2000 |
| Předmět: |
Male
Glycosylation Molecular Sequence Data Disaccharide Hyaluronoglucosaminidase Oligosaccharides Disaccharides Biochemistry Mass Spectrometry Dermatan sulfate Substrate Specificity Glycosaminoglycan chemistry.chemical_compound Sulfation Hyaluronidase Testis Hyaluronic acid medicine Animals Chondroitin Chondroitin sulfate Molecular Biology Chromatography High Pressure Liquid Glycosaminoglycans Sulfates Chondroitin Sulfates Streptococcus General Medicine carbohydrates (lipids) Carbohydrate Sequence chemistry Cattle medicine.drug |
| Zdroj: | Journal of Biochemistry. 127:695-702 |
| ISSN: | 0021-924X |
| DOI: | 10.1093/oxfordjournals.jbchem.a022659 |
| Popis: | A method was developed for the reconstruction of glycosaminoglycan (GAG) oligosaccharides using the transglycosylation reaction of an endo-beta-N-acetylhexosaminidase, testicular hyaluronidase, under optimal conditions. Repetition of the transglycosylation using suitable combinations of various GAGs as acceptors and donors made it possible to custom-synthesize GAG oligosaccharides. Thus we prepared a library of chimeric GAG oligosaccharides with hybrid structures composed of disaccharide units such as GlcA-GlcNAc (from hyaluronic acid), GlcA-GalNAc (from chondroitin), GlcA-GalNAc4S (from chondroitin 4-sulfate), GlcA-GalNAc6S (from chondroitin 6-sulfate), IdoA-GalNAc (from desulfated dermatan sulfate), and GlcA-GalNAc4,6-diS (from chondroitin sulfate E). The specificity of the hyaluronidase from Streptococcus dysgalactiae (hyaluronidase SD) was then investigated using these chimeric GAG oligosaccharides as model substrates. The results indicate that the specificity of hyaluronidase SD is determined by the following restrictions at the nonreducing terminal side of the cleavage site: (i) at least one disaccharide unit (GlcA-GlcNAc) is necessary for the enzymatic action of hyaluronidase SD; (ii) cleavage is inhibited by sulfation of the N-acetylgalactosamine; (iii) hyaluronidase SD releases GlcA-GalNAc and IdoA-GalNAc units as well as GlcA-GlcNAc. At the reducing terminal side of the cleavage site, the sulfated residues on the N-acetylgalactosamines in the disaccharide units were found to have no influence on the cleavage. Additionally, we found that hyaluronidase SD can specifically and endolytically cleave the internal unsulfated regions of chondroitin sulfate chains. This demonstration indicates that custom-synthesized GAG oligosaccharides will open a new avenue in GAG glycotechnology. |
| Databáze: | OpenAIRE |
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