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Purification and Characterization of a Dark Red Skin Related Dimeric Polyphenol Oxidase from Huaniu Apples

The distinct dark-red skin of Huaniu apples renders them attractive to customers. However, the mechanism that leads to the development of the color of the fruit is unclear. In this study, we found that compared with red Fuji (a bright-red apple cultivar), Huaniu apples had higher contents of (−)-epi...

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Autores principales: Liu, Bin, Zhou, Xianfang, Guan, Haiyan, Pang, Xuequn, Zhang, Zhaoqi
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9223062/
https://www.ncbi.nlm.nih.gov/pubmed/35741987
http://dx.doi.org/10.3390/foods11121790
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author Liu, Bin
Zhou, Xianfang
Guan, Haiyan
Pang, Xuequn
Zhang, Zhaoqi
author_facet Liu, Bin
Zhou, Xianfang
Guan, Haiyan
Pang, Xuequn
Zhang, Zhaoqi
author_sort Liu, Bin
collection PubMed
description The distinct dark-red skin of Huaniu apples renders them attractive to customers. However, the mechanism that leads to the development of the color of the fruit is unclear. In this study, we found that compared with red Fuji (a bright-red apple cultivar), Huaniu apples had higher contents of (−)-epicatechin (EC), (−)-epigallocatechin (EGC), (−)-gallocatechin gallate (GCG), and procyanidins (PCs) B2 and C1 in the peel, which implies that the polymerization of the flavanols and PCs may be correlated with the dark-red skin of the fruit. Using EC as a substrate, we purified an enzyme from Huaniu peel. We performed protein sequencing and discovered that the enzyme was a polyphenol oxidase (PPO). The molecular weight of the enzyme was approximately 140 kDa, which we estimated by native-PAGE and SDS-PAGE, while it was 61 kDa by urea-SDS-PAGE, from which we discovered that the PPO was a dimer. We observed the lowest K(m) value for catechol (0.60 mM), and the best substrate was 4-methylcatechol, with a V(max) of 526.32 U mg(−1) protein. EC is a suitable natural substrate, with a K(m) value of 1.17 mM, and 55.27% of the V(max)/K(m) of 4-methylcatechol. When we used EC as a substrate, the optimum temperature and pH of the PPO were 25 °C and 5.0, respectively. In summary, we purified a dimeric PPO from Huaniu apples that showed high activity to EC, which might catalyze the polymerization of flavanols and PCs and lead to the dark-red color development of the fruit.
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spelling pubmed-92230622022-06-24 Purification and Characterization of a Dark Red Skin Related Dimeric Polyphenol Oxidase from Huaniu Apples Liu, Bin Zhou, Xianfang Guan, Haiyan Pang, Xuequn Zhang, Zhaoqi Foods Article The distinct dark-red skin of Huaniu apples renders them attractive to customers. However, the mechanism that leads to the development of the color of the fruit is unclear. In this study, we found that compared with red Fuji (a bright-red apple cultivar), Huaniu apples had higher contents of (−)-epicatechin (EC), (−)-epigallocatechin (EGC), (−)-gallocatechin gallate (GCG), and procyanidins (PCs) B2 and C1 in the peel, which implies that the polymerization of the flavanols and PCs may be correlated with the dark-red skin of the fruit. Using EC as a substrate, we purified an enzyme from Huaniu peel. We performed protein sequencing and discovered that the enzyme was a polyphenol oxidase (PPO). The molecular weight of the enzyme was approximately 140 kDa, which we estimated by native-PAGE and SDS-PAGE, while it was 61 kDa by urea-SDS-PAGE, from which we discovered that the PPO was a dimer. We observed the lowest K(m) value for catechol (0.60 mM), and the best substrate was 4-methylcatechol, with a V(max) of 526.32 U mg(−1) protein. EC is a suitable natural substrate, with a K(m) value of 1.17 mM, and 55.27% of the V(max)/K(m) of 4-methylcatechol. When we used EC as a substrate, the optimum temperature and pH of the PPO were 25 °C and 5.0, respectively. In summary, we purified a dimeric PPO from Huaniu apples that showed high activity to EC, which might catalyze the polymerization of flavanols and PCs and lead to the dark-red color development of the fruit. MDPI 2022-06-17 /pmc/articles/PMC9223062/ /pubmed/35741987 http://dx.doi.org/10.3390/foods11121790 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Liu, Bin
Zhou, Xianfang
Guan, Haiyan
Pang, Xuequn
Zhang, Zhaoqi
Purification and Characterization of a Dark Red Skin Related Dimeric Polyphenol Oxidase from Huaniu Apples
title Purification and Characterization of a Dark Red Skin Related Dimeric Polyphenol Oxidase from Huaniu Apples
title_full Purification and Characterization of a Dark Red Skin Related Dimeric Polyphenol Oxidase from Huaniu Apples
title_fullStr Purification and Characterization of a Dark Red Skin Related Dimeric Polyphenol Oxidase from Huaniu Apples
title_full_unstemmed Purification and Characterization of a Dark Red Skin Related Dimeric Polyphenol Oxidase from Huaniu Apples
title_short Purification and Characterization of a Dark Red Skin Related Dimeric Polyphenol Oxidase from Huaniu Apples
title_sort purification and characterization of a dark red skin related dimeric polyphenol oxidase from huaniu apples
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9223062/
https://www.ncbi.nlm.nih.gov/pubmed/35741987
http://dx.doi.org/10.3390/foods11121790
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