title: Structural understanding of the recycling of oxidized ascorbate by dehydroascorbate reductase (OsDHAR) from Oryza sativa L. japonica.
abstract: Dehydroascorbate reductase (DHAR) is a key enzyme involved in the recycling of ascorbate, which catalyses the glutathione (GSH)-dependent reduction of oxidized ascorbate (dehydroascorbate, DHA). As a result, DHAR regenerates a pool of reduced ascorbate and detoxifies reactive oxygen species (ROS). In previous experiments involving transgenic rice, we observed that overexpression of DHAR enhanced grain yield and biomass. Since the structure of DHAR is not available, the enzymatic mechanism is not well-understood and remains poorly characterized. To elucidate the molecular basis of DHAR catalysis, we determined the crystal structures of DHAR from Oryza sativa L. japonica (OsDHAR) in the native, ascorbate-bound, and GSH-bound forms and refined their resolutions to 1.9, 1.7, and 1.7 Å, respectively. These complex structures provide the first information regarding the location of the ascorbate and GSH binding sites and their interacting residues. The location of the ascorbate-binding site overlaps with the GSH-binding site, suggesting a ping-pong kinetic mechanism for electron transfer at the common Cys20 active site. Our structural information and mutagenesis data provide useful insights into the reaction mechanism of OsDHAR against ROS-induced oxidative stress in rice.
authors.affiliation: Division of Polar Life Sciences, Korea Polar Research Institute, Incheon 406-840, Republic of Korea.
authors.forename: Hackwon
authors.initials: H
authors.lastname: Do
authors.affiliation: School of Life Sciences, BK21 Plus KNU Creative BioResearch Group, Kyungpook National University, Daegu 702-701, Republic of Korea.
authors.forename: Il-Sup
authors.initials: IS
authors.lastname: Kim
authors.affiliation: Division of Polar Life Sciences, Korea Polar Research Institute, Incheon 406-840, Republic of Korea.
authors.forename: Byoung Wook
authors.initials: BW
authors.lastname: Jeon
authors.affiliation: Division of Polar Life Sciences, Korea Polar Research Institute, Incheon 406-840, Republic of Korea.|Department of Polar Sciences, University of Science and Technology, Incheon 406-840, Republic of Korea.
authors.forename: Chang Woo
authors.initials: CW
authors.lastname: Lee
authors.affiliation: Division of Polar Life Sciences, Korea Polar Research Institute, Incheon 406-840, Republic of Korea.
authors.forename: Ae Kyung
authors.initials: AK
authors.lastname: Park
authors.affiliation: Division of Polar Life Sciences, Korea Polar Research Institute, Incheon 406-840, Republic of Korea.
authors.forename: Ah Ram
authors.initials: AR
authors.lastname: Wi
authors.affiliation: Division of Polar Life Sciences, Korea Polar Research Institute, Incheon 406-840, Republic of Korea.
authors.forename: Seung Chul
authors.initials: SC
authors.lastname: Shin
authors.affiliation: Division of Polar Life Sciences, Korea Polar Research Institute, Incheon 406-840, Republic of Korea.|Department of Polar Sciences, University of Science and Technology, Incheon 406-840, Republic of Korea.
authors.forename: Hyun
authors.initials: H
authors.lastname: Park
authors.affiliation: Division of Biological Sciences, University of California San Diego, La Jolla, California 92093-0116, USA.
authors.forename: Young-Saeng
authors.initials: YS
authors.lastname: Kim
authors.affiliation: School of Life Sciences, BK21 Plus KNU Creative BioResearch Group, Kyungpook National University, Daegu 702-701, Republic of Korea.
authors.forename: Ho-Sung
authors.initials: HS
authors.lastname: Yoon
authors.affiliation: Division of Polar Life Sciences, Korea Polar Research Institute, Incheon 406-840, Republic of Korea.|Department of Polar Sciences, University of Science and Technology, Incheon 406-840, Republic of Korea.
authors.forename: Han-Woo
authors.initials: HW
authors.lastname: Kim
authors.affiliation: Division of Polar Life Sciences, Korea Polar Research Institute, Incheon 406-840, Republic of Korea.|Department of Polar Sciences, University of Science and Technology, Incheon 406-840, Republic of Korea.
authors.forename: Jun Hyuck
authors.initials: JH
authors.lastname: Lee
mesh_terms: D000595:Amino Acid Sequence; D001205:Ascorbic Acid / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D020134:Catalytic Domain; D015870:Gene Expression; D005978:Glutathione / Q000737:chemistry / Q000235:genetics; D053858:Metabolic Networks and Pathways; D008958:Models, Molecular; D009154:Mutation; D012275:Oryza / Q000378:metabolism*; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D017382:Reactive Oxygen Species / Q000378:metabolism; D013312:Stress, Physiological
chemical_list: D017382:Reactive Oxygen Species; D010088:Oxidoreductases; C020666:glutathione dehydrogenase (ascorbate); D005978:Glutathione; D001205:Ascorbic Acid