title: Molecular cloning, substrate specificity of the functionally expressed dihydroflavonol 4-reductases from Malus domestica and Pyrus communis cultivars and the consequences for flavonoid metabolism.
abstract: Treatment with the dioxygenase inhibitor prohexadione-Ca leads to major changes in the flavonoid metabolism of apple (Malus domestica) and pear (Pyrus communis) leaves. Accumulation of unusual 3-deoxyflavonoids is observed, which have been linked to an enhanced resistance toward fire blight. The committed step in this pathway is the reduction of flavanones. Crude extracts from leaves are able to perform this reaction. There was previous evidence that DFR enzymes of certain plants possess additional flavanone 4-reductase (FNR) activity. Such an FNR activity of DFR enzymes is proved here by heterologous expression of the enzymes. The heterologously expressed DFR/FNR enzymes of Malus and Pyrus possess distinct differences in substrate specificities despite only minor differences of the amino acid sequences. Kinetic studies showed that dihydroflavonols generally are the preferred substrates. However, with the observed substrate specificities the occurrence of 3-deoxyflavonoids in vivo after application of prohexadione-Ca can be explained.
authors.affiliation: Chair of Floriculture Crops and Horticultural Plant Breeding, TUM Weihenstephan, Am Hochanger 4, D-85350 Freising, Germany.
authors.forename: Thilo C
authors.initials: TC
authors.lastname: Fischer
authors.forename: Heidrun
authors.initials: H
authors.lastname: Halbwirth
authors.forename: Barbara
authors.initials: B
authors.lastname: Meisel
authors.forename: Karl
authors.initials: K
authors.lastname: Stich
authors.forename: Gert
authors.initials: G
authors.lastname: Forkmann
mesh_terms: D000429:Alcohol Oxidoreductases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D018744:DNA, Plant / Q000235:genetics; D005419:Flavonoids / Q000378:metabolism; D017343:Genes, Plant; D007700:Kinetics; D027845:Malus / Q000201:enzymology* / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D018515:Plant Leaves / Q000201:enzymology; D031989:Pyrus / Q000201:enzymology* / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
chemical_list: D018076:DNA, Complementary; D018744:DNA, Plant; D005419:Flavonoids; D011994:Recombinant Proteins; D000429:Alcohol Oxidoreductases; C056570:dihydroflavanol 4-reductase