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Structure and catalytic mechanism of monodehydroascorbate reductase, MDHAR, from Oryza sativa L. japonica

Ascorbic acid (AsA) maintains redox homeostasis by scavenging reactive oxygen species from prokaryotes to eukaryotes, especially plants. The enzyme monodehydroascorbate reductase (MDHAR) regenerates AsA by catalysing the reduction of monodehydroascorbate, using NADH or NADPH as an electron donor. Th...

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Autores principales: Park, Ae Kyung, Kim, Il-Sup, Do, Hackwon, Jeon, Byung Wook, Lee, Chang Woo, Roh, Soo Jung, Shin, Seung Chul, Park, Hyun, Kim, Young-Saeng, Kim, Yul-Ho, Yoon, Ho-Sung, Lee, Jun Hyuck, Kim, Han-Woo
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5031999/
https://www.ncbi.nlm.nih.gov/pubmed/27652777
http://dx.doi.org/10.1038/srep33903
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author Park, Ae Kyung
Kim, Il-Sup
Do, Hackwon
Jeon, Byung Wook
Lee, Chang Woo
Roh, Soo Jung
Shin, Seung Chul
Park, Hyun
Kim, Young-Saeng
Kim, Yul-Ho
Yoon, Ho-Sung
Lee, Jun Hyuck
Kim, Han-Woo
author_facet Park, Ae Kyung
Kim, Il-Sup
Do, Hackwon
Jeon, Byung Wook
Lee, Chang Woo
Roh, Soo Jung
Shin, Seung Chul
Park, Hyun
Kim, Young-Saeng
Kim, Yul-Ho
Yoon, Ho-Sung
Lee, Jun Hyuck
Kim, Han-Woo
author_sort Park, Ae Kyung
collection PubMed
description Ascorbic acid (AsA) maintains redox homeostasis by scavenging reactive oxygen species from prokaryotes to eukaryotes, especially plants. The enzyme monodehydroascorbate reductase (MDHAR) regenerates AsA by catalysing the reduction of monodehydroascorbate, using NADH or NADPH as an electron donor. The detailed recycling mechanism of MDHAR remains unclear due to lack of structural information. Here, we present the crystal structures of MDHAR in the presence of cofactors, nicotinamide adenine dinucleotide (NAD(+)) and nicotinamide adenine dinucleotide phosphate (NADP(+)), and complexed with AsA as well as its analogue, isoascorbic acid (ISD). The overall structure of MDHAR is similar to other iron-sulphur protein reductases, except for a unique long loop of 63–80 residues, which seems to be essential in forming the active site pocket. From the structural analysis and structure-guided point mutations, we found that the Arg320 residue plays a major substrate binding role, and the Tyr349 residue mediates electron transfer from NAD(P)H to bound substrate via FAD. The enzymatic activity of MDHAR favours NADH as an electron donor over NADPH. Our results show, for the first time, structural insights into this preference. The MDHAR-ISD complex structure revealed an alternative binding conformation of ISD, compared with the MDHAR-AsA complex. This implies a broad substrate (antioxidant) specificity and resulting greater protective ability of MDHAR.
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spelling pubmed-50319992016-09-29 Structure and catalytic mechanism of monodehydroascorbate reductase, MDHAR, from Oryza sativa L. japonica Park, Ae Kyung Kim, Il-Sup Do, Hackwon Jeon, Byung Wook Lee, Chang Woo Roh, Soo Jung Shin, Seung Chul Park, Hyun Kim, Young-Saeng Kim, Yul-Ho Yoon, Ho-Sung Lee, Jun Hyuck Kim, Han-Woo Sci Rep Article Ascorbic acid (AsA) maintains redox homeostasis by scavenging reactive oxygen species from prokaryotes to eukaryotes, especially plants. The enzyme monodehydroascorbate reductase (MDHAR) regenerates AsA by catalysing the reduction of monodehydroascorbate, using NADH or NADPH as an electron donor. The detailed recycling mechanism of MDHAR remains unclear due to lack of structural information. Here, we present the crystal structures of MDHAR in the presence of cofactors, nicotinamide adenine dinucleotide (NAD(+)) and nicotinamide adenine dinucleotide phosphate (NADP(+)), and complexed with AsA as well as its analogue, isoascorbic acid (ISD). The overall structure of MDHAR is similar to other iron-sulphur protein reductases, except for a unique long loop of 63–80 residues, which seems to be essential in forming the active site pocket. From the structural analysis and structure-guided point mutations, we found that the Arg320 residue plays a major substrate binding role, and the Tyr349 residue mediates electron transfer from NAD(P)H to bound substrate via FAD. The enzymatic activity of MDHAR favours NADH as an electron donor over NADPH. Our results show, for the first time, structural insights into this preference. The MDHAR-ISD complex structure revealed an alternative binding conformation of ISD, compared with the MDHAR-AsA complex. This implies a broad substrate (antioxidant) specificity and resulting greater protective ability of MDHAR. Nature Publishing Group 2016-09-22 /pmc/articles/PMC5031999/ /pubmed/27652777 http://dx.doi.org/10.1038/srep33903 Text en Copyright © 2016, The Author(s) http://creativecommons.org/licenses/by/4.0/ This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/
spellingShingle Article
Park, Ae Kyung
Kim, Il-Sup
Do, Hackwon
Jeon, Byung Wook
Lee, Chang Woo
Roh, Soo Jung
Shin, Seung Chul
Park, Hyun
Kim, Young-Saeng
Kim, Yul-Ho
Yoon, Ho-Sung
Lee, Jun Hyuck
Kim, Han-Woo
Structure and catalytic mechanism of monodehydroascorbate reductase, MDHAR, from Oryza sativa L. japonica
title Structure and catalytic mechanism of monodehydroascorbate reductase, MDHAR, from Oryza sativa L. japonica
title_full Structure and catalytic mechanism of monodehydroascorbate reductase, MDHAR, from Oryza sativa L. japonica
title_fullStr Structure and catalytic mechanism of monodehydroascorbate reductase, MDHAR, from Oryza sativa L. japonica
title_full_unstemmed Structure and catalytic mechanism of monodehydroascorbate reductase, MDHAR, from Oryza sativa L. japonica
title_short Structure and catalytic mechanism of monodehydroascorbate reductase, MDHAR, from Oryza sativa L. japonica
title_sort structure and catalytic mechanism of monodehydroascorbate reductase, mdhar, from oryza sativa l. japonica
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5031999/
https://www.ncbi.nlm.nih.gov/pubmed/27652777
http://dx.doi.org/10.1038/srep33903
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