title: Sequential enzymatic glycosyltransfer reactions on a microfluidic device: Synthesis of a glycosaminoglycan linkage region tetrasaccharide.
abstract: A microfluidic chip carrying three reaction chambers was designed and constructed to examine sequential multiple enzymatic reactions. The synthesis of oligosaccharides in living cells is carried out in the Golgi apparatus where multiple enzymes such as glycosidase and glycosyltransferases act on a variety of substrates to generate glycoconjugates that include glycolipids and glycoproteins. The regulatory mechanism of the process however remains unknown. A microchip-based analysis platform may provide a valuable tool with which to address the issue by mimicking the Golgi function. We thus examined 3 sequential glycosyltransfer reactions on a chip, and succeeded in the synthesis of a tetrasaccharide using immobilized enzymes. Also, the kinetic parameters for a recently identified glycosyltransferase, proteoglycan GalT-I, were obtained for the first time.
authors.affiliation: Mitsubishi Kagaku Institute of Life Sciences (MITILS), 11 Minamiooya, Machida-shi, Tokyo, 194-8511, Japan.
authors.forename: Yasunari
authors.initials: Y
authors.lastname: Ono
authors.forename: Motohiro
authors.initials: M
authors.lastname: Kitajima
authors.forename: Shusaku
authors.initials: S
authors.lastname: Daikoku
authors.forename: Toshifumi
authors.initials: T
authors.lastname: Shiroya
authors.forename: Shoko
authors.initials: S
authors.lastname: Nishihara
authors.forename: Yoshimi
authors.initials: Y
authors.lastname: Kanie
authors.forename: Katsuhiko
authors.initials: K
authors.lastname: Suzuki
authors.forename: Satoshi
authors.initials: S
authors.lastname: Goto
authors.forename: Osamu
authors.initials: O
authors.lastname: Kanie
mesh_terms: D002240:Carbohydrate Sequence; D004800:Enzymes, Immobilized; D008401:Gas Chromatography-Mass Spectrometry; D006025:Glycosaminoglycans / Q000737:chemistry* / Q000378:metabolism; D006031:Glycosylation; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000706:statistics & numerical data*; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000138:chemical synthesis* / Q000737:chemistry; D012685:Sepharose / Q000737:chemistry
chemical_list: D004800:Enzymes, Immobilized; D006025:Glycosaminoglycans; D009844:Oligosaccharides; D012685:Sepharose