1. The structural and functional characterization of Malus domestica double bond reductase MdDBR provides insights towards the identification of its substrates.
- Author
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Caliandro R, Polsinelli I, Demitri N, Musiani F, Martens S, and Benini S
- Subjects
- Amino Acid Sequence, Apoproteins genetics, Apoproteins metabolism, Arabidopsis chemistry, Arabidopsis enzymology, Binding Sites, Butanones metabolism, Cloning, Molecular, Crystallography, X-Ray, Escherichia coli genetics, Escherichia coli metabolism, Gene Expression, Genetic Vectors chemistry, Genetic Vectors metabolism, Kinetics, Malus enzymology, Models, Molecular, NADP metabolism, Oxidoreductases genetics, Oxidoreductases metabolism, Plant Proteins genetics, Plant Proteins metabolism, Protein Binding, Protein Conformation, alpha-Helical, Protein Conformation, beta-Strand, Protein Interaction Domains and Motifs, Recombinant Proteins chemistry, Recombinant Proteins genetics, Recombinant Proteins metabolism, Rubus chemistry, Rubus enzymology, Sequence Alignment, Sequence Homology, Amino Acid, Substrate Specificity, Thermodynamics, Nicotiana chemistry, Nicotiana enzymology, Apoproteins chemistry, Butanones chemistry, Malus chemistry, NADP chemistry, Oxidoreductases chemistry, Plant Proteins chemistry
- Abstract
In this study we describe the crystal structures of the apoform, the binary and the ternary complexes of a double bond reductase from Malus domestica L. (MdDBR) and explore a range of potential substrates. The overall fold of MdDBR is similar to that of the medium chain reductase/dehydrogenase/zinc-dependent alcohol dehydrogenase-like family. Structural comparison of MdDBR with Arabidopsis thaliana DBR (AtDBR), Nicotiana tabacum DBR (NtDBR) and Rubus idaeus DBR (RiDBR) allowed the identification of key amino acids involved in cofactor and ligands binding and shed light on how these residues may guide the orientation of the substrates. The enzyme kinetic for the substrate trans-4-phenylbuten-2-one has been analyzed, and MdDBR activity towards a variety of substrates was tested. This enzyme has been reported to be involved in the phenylpropanoid pathway where it would catalyze the NADPH-dependent reduction of the α, β-unsaturated double bond of carbonyl metabolites. Our study provides new data towards the identification of MdDBR natural substrate and the biosynthetic pathway where it belongs. Furthermore, the originally proposed involvement in dihydrochalcone biosynthesis in apple must be questioned., (Copyright © 2020 Elsevier B.V. All rights reserved.)
- Published
- 2021
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