title: Synthesis of Glc1Man9-Glycoprotein Probes by a Misfolding/Enzymatic Glucosylation/Misfolding Sequence.
abstract: Glycoproteins in non-native conformations are often toxic to cells and may cause diseases, thus the quality control (QC) system eliminates these unwanted species. Lectin chaperone calreticulin and glucosidase II, both of which recognize the Glc1 Man9 oligosaccharide on glycoproteins, are important components of the glycoprotein QC system. Reported herein is the preparation of Glc1 Man9 -glycoproteins in both native and non-native conformations by using the following sequence: misfolding of chemically synthesized Man9 -glycoprotein, enzymatic glucosylation, and another misfolding step. By using synthetic glycoprotein probes, calreticulin was found to bind preferentially to a hydrophobic non-native glycoprotein whereas glucosidase II activity was not affected by glycoprotein conformation. The results demonstrate the ability of chemical synthesis to deliver homogeneous glycoproteins in several non-native conformations for probing the glycoprotein QC system.
authors.affiliation: Department of Chemistry, Graduate School of Science, Osaka University, 1-1 Machikaneyama, Toyonaka, Osaka, 560-0043, Japan.
authors.forename: Masayuki
authors.identifier: http://orcid.org/0000-0001-6486-9678
authors.initials: M
authors.lastname: Izumi
authors.affiliation: Department of Chemistry, Graduate School of Science, Osaka University, 1-1 Machikaneyama, Toyonaka, Osaka, 560-0043, Japan.
authors.forename: Yukiho
authors.initials: Y
authors.lastname: Oka
authors.affiliation: Department of Chemistry, Graduate School of Science, Osaka University, 1-1 Machikaneyama, Toyonaka, Osaka, 560-0043, Japan.
authors.forename: Ryo
authors.initials: R
authors.lastname: Okamoto
authors.affiliation: ERATO Ito glycotrilogy project Japan Science and Technology Agency (JST), 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan.
authors.forename: Akira
authors.initials: A
authors.lastname: Seko
authors.affiliation: ERATO Ito glycotrilogy project Japan Science and Technology Agency (JST), 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan.
authors.forename: Yoichi
authors.initials: Y
authors.lastname: Takeda
authors.affiliation: ERATO Ito glycotrilogy project Japan Science and Technology Agency (JST), 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan. yukito@riken.jp.|Synthetic Cellular Chemistry Laboratory, RIKEN, 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan. yukito@riken.jp.
authors.forename: Yukishige
authors.initials: Y
authors.lastname: Ito
authors.affiliation: Department of Chemistry, Graduate School of Science, Osaka University, 1-1 Machikaneyama, Toyonaka, Osaka, 560-0043, Japan. kajihara@chem.sci.osaka-u.ac.jp.|ERATO Ito glycotrilogy project Japan Science and Technology Agency (JST), 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan. kajihara@chem.sci.osaka-u.ac.jp.
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Kajihara
mesh_terms: D002851:Chromatography, High Pressure Liquid; D005964:Glucosyltransferases / Q000737:chemistry*; D006023:Glycoproteins / Q000737:chemistry*; D015335:Molecular Probes / Q000138:chemical synthesis*
chemical_list: D006023:Glycoproteins; D015335:Molecular Probes; D005964:Glucosyltransferases
keywords: enzymes; glycopeptides; glycoproteins; mass spectrometry; protein folding