Biochemical and Functional Characterization of Anthocyanidin Reductase (ANR) from <i>Mangifera indica</i> L.
Lin Tan,
Mei Wang,
Youfa Kang,
Farrukh Azeem,
Zhaoxi Zhou,
Decai Tuo,
Lina María Preciado Rojo,
Ikhlas A. Khan,
Zhiqiang Pan
Affiliations
Lin Tan
Haikou Experimental Station, Chinese Academy of Tropical Agricultural Sciences (CATAS)—Hainan Key Laboratory of Banana Genetic Improvement, Haikou 570102, Hainan Province, China
Mei Wang
National Center for Natural Products Research, Thad Cochran Research Center, School of Pharmacy, University of Mississippi, University, MS 38677, USA
Youfa Kang
Department of Basic Education in Liberal Arts, Hainan University, Haikou 571101, Hainan Province, China
Farrukh Azeem
Haikou Experimental Station, Chinese Academy of Tropical Agricultural Sciences (CATAS)—Hainan Key Laboratory of Banana Genetic Improvement, Haikou 570102, Hainan Province, China
Zhaoxi Zhou
Haikou Experimental Station, Chinese Academy of Tropical Agricultural Sciences (CATAS)—Hainan Key Laboratory of Banana Genetic Improvement, Haikou 570102, Hainan Province, China
Decai Tuo
Institute of Tropical Bioscience and Biotechnology, Chinese Academy of Tropical Agricultural Sciences (CATAS), Haikou 571101, Hainan Province, China
Lina María Preciado Rojo
Programa Ofidismo-Escorpionismo, Facultad de Ciencias Farmacéuticas y Alimentarias, Universidad de Antioquia, Medellín 1226, Colombia
Ikhlas A. Khan
National Center for Natural Products Research, Thad Cochran Research Center, School of Pharmacy, University of Mississippi, University, MS 38677, USA
Zhiqiang Pan
United States Department of Agriculture, Agricultural Research Service, Natural Products Utilization Research Unit (USDA-ARS-NPURU), P.O. Box 1848, University, MS 38677-1848, USA
Mango (Mangifera indica L.) is abundant in proanthocyanidins (PAs) that are important for human health and plant response to abiotic stresses. However, the molecular mechanisms involved in PA biosynthesis still need to be elucidated. Anthocyanidin reductase (ANR) catalyzes a key step in PA biosynthesis. In this study, three ANR cDNAs (MiANR1-1,1-2,1-3) were isolated from mango, and expressed in Escherichia coli. In vitro enzyme assay showed MiANR proteins convert cyanidin to their corresponding flavan-3-ols, such as (−)-catechin and (−)-epicatechin. Despite high amino acid similarity, the recombinant ANR proteins exhibited differences in enzyme kinetics and cosubstrate preference. MiANR1-2 and MiANR1-3 have the same optimum pH of 4.0 in citrate buffer, while the optimum pH for MiANR1-1 is pH 3.0 in phosphate buffer. MiANR1-1 does not use either NADPH or NADH as co-substrate while MiANR1-2/1-3 use only NADPH as co-substrate. MiANR1-2 has the highest Km and Vmax for cyanidin, followed by MiANR1-3 and MiANR1-1. The overexpression of MiANRs in ban mutant reconstructed the biosynthetic pathway of PAs in the seed coat. These data demonstrate MiANRs can form the ANR pathway, leading to the formation of two types of isomeric flavan-3-ols and PAs in mango.
