title: Structural and functional study of D-glucuronyl C5-epimerase.
abstract: Heparan sulfate (HS) is a glycosaminoglycan present on the cell surface and in the extracellular matrix, which interacts with diverse signal molecules and is essential for many physiological processes including embryonic development, cell growth, inflammation, and blood coagulation. D-glucuronyl C5-epimerase (Glce) is a crucial enzyme in HS synthesis, converting D-glucuronic acid to L-iduronic acid to increase HS flexibility. This modification of HS is important for protein ligand recognition. We have determined the crystal structures of Glce in apo-form (unliganded) and in complex with heparin hexasaccharide (product of Glce following O-sulfation), both in a stable dimer conformation. A Glce dimer contains two catalytic sites, each at a positively charged cleft in C-terminal α-helical domains binding one negatively charged hexasaccharide. Based on the structural and mutagenesis studies, three tyrosine residues, Tyr(468), Tyr(528), and Tyr(546), in the active site were found to be crucial for the enzymatic activity. The complex structure also reveals the mechanism of product inhibition (i.e. 2-O- and 6-O-sulfation of HS keeps the C5 carbon of L-iduronic acid away from the active-site tyrosine residues). Our structural and functional data advance understanding of the key modification in HS biosynthesis.
authors.affiliation: From the Glycochemistry and Glycobiology Laboratory, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zu Chong Zhi Road, Pudong, Shanghai 201203, China,; the VARI-SIMM Center, Center for Structure and Function of Drug Targets, Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Qin
authors.affiliation: Laboratory of Structural Sciences, Center for Structural Biology and Drug Discovery, Van Andel Research Institute, Grand Rapids, Michigan 49503,. Electronic address: Jiyuan.Ke@vai.org.
authors.forename: Jiyuan
authors.initials: J
authors.lastname: Ke
authors.affiliation: Laboratory of Structural Sciences, Center for Structural Biology and Drug Discovery, Van Andel Research Institute, Grand Rapids, Michigan 49503.
authors.forename: Xin
authors.initials: X
authors.lastname: Gu
authors.affiliation: From the Glycochemistry and Glycobiology Laboratory, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zu Chong Zhi Road, Pudong, Shanghai 201203, China,; the Department of Medical Biochemistry and Microbiology, University of Uppsala, Biomedical Center, SE-751 23 Uppsala, Sweden, and.
authors.forename: Jianping
authors.initials: J
authors.lastname: Fang
authors.affiliation: From the Glycochemistry and Glycobiology Laboratory, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zu Chong Zhi Road, Pudong, Shanghai 201203, China.
authors.forename: Wucheng
authors.initials: W
authors.lastname: Wang
authors.affiliation: From the Glycochemistry and Glycobiology Laboratory, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zu Chong Zhi Road, Pudong, Shanghai 201203, China.
authors.forename: Qifei
authors.initials: Q
authors.lastname: Cong
authors.affiliation: From the Glycochemistry and Glycobiology Laboratory, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zu Chong Zhi Road, Pudong, Shanghai 201203, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Li
authors.affiliation: the VARI-SIMM Center, Center for Structure and Function of Drug Targets, Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.
authors.forename: Jinzhi
authors.initials: J
authors.lastname: Tan
authors.affiliation: the Life Sciences Collaborative Access Team, Synchrotron Research Center, Northwestern University, Argonne, Illinois 60439.
authors.forename: Joseph S
authors.initials: JS
authors.lastname: Brunzelle
authors.affiliation: the VARI-SIMM Center, Center for Structure and Function of Drug Targets, Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.
authors.forename: Chenghai
authors.initials: C
authors.lastname: Zhang
authors.affiliation: the VARI-SIMM Center, Center for Structure and Function of Drug Targets, Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Jiang
authors.affiliation: Laboratory of Structural Sciences, Center for Structural Biology and Drug Discovery, Van Andel Research Institute, Grand Rapids, Michigan 49503.
authors.forename: Karsten
authors.initials: K
authors.lastname: Melcher
authors.affiliation: the Department of Medical Biochemistry and Microbiology, University of Uppsala, Biomedical Center, SE-751 23 Uppsala, Sweden, and.
authors.forename: Jin-Ping
authors.initials: JP
authors.lastname: Li
authors.affiliation: the VARI-SIMM Center, Center for Structure and Function of Drug Targets, Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China,; Laboratory of Structural Sciences, Center for Structural Biology and Drug Discovery, Van Andel Research Institute, Grand Rapids, Michigan 49503,. Electronic address: Eric.Xu@vai.org.
authors.forename: H Eric
authors.initials: HE
authors.lastname: Xu
authors.affiliation: From the Glycochemistry and Glycobiology Laboratory, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zu Chong Zhi Road, Pudong, Shanghai 201203, China,. Electronic address: dingkan@simm.ac.cn.
authors.forename: Kan
authors.initials: K
authors.lastname: Ding
mesh_terms: D000818:Animals; D002238:Carbohydrate Epimerases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D018360:Crystallography, X-Ray; D006497:Heparan Sulfate / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D015027:Zebrafish; D029961:Zebrafish Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D002238:Carbohydrate Epimerases; D006497:Heparan Sulfate; D029961:Zebrafish Proteins
keywords: Crystal Structure; Enzyme Mechanism; Epimerization; Glce; Glycosaminoglycan; Heparan Sulfate; Heparin; d-Glucuronyl C5 Epimerase