Characterization and functional identification of a novel plant 4,5-extradiol dioxygenase involved in betalain pigment biosynthesis in Portulaca grandiflora.
Betalains are pigments that replace anthocyanins in the majority of families of the plant order Caryophyllales. Betalamic acid is the common chromophore of betalains. The key enzyme of the betalain biosynthetic pathway is an extradiol dioxygenase that opens the cyclic ring of dihydroxy-phenylalanine (DOPA) between carbons 4 and 5, thus producing an unstable seco-DOPA that rearranges nonenzymatically to betalamic acid. A gene for a 4,5-DOPA-dioxygenase has already been isolated from the fungus Amanita muscaria, but no homolog was ever found in plants. To identify the plant gene, we constructed subtractive libraries between different colored phenotypes of isogenic lines of Portulaca grandiflora (Portulacaceae) and between different stages of flower bud formation. Using in silico analysis of differentially expressed cDNAs, we identified a candidate showing strong homology at the level of translated protein with the LigB domain present in several bacterial extradiol 4,5-dioxygenases. The gene was expressed only in colored flower petals. The function of this gene in the betalain biosynthetic pathway was confirmed by biolistic genetic complementation in white petals of P. grandiflora genotypes lacking the gene for color formation. This gene named DODA is the first characterized member of a novel family of plant dioxygenases phylogenetically distinct from Amanita sp. DOPA-dioxygenase. Homologs of DODA are present not only in betalain-producing plants but also, albeit with some changes near the catalytic site, in other angiosperms and in the bryophyte Physcomitrella patens. These homologs are part of a novel conserved plant gene family probably involved in aromatic compound metabolism.
Christinet
Laurent
L
Laboratory of Plant Cell Genetics, Department of Plant Molecular Biology, University of Lausanne, CH 1015 Lausanne, Switzerland.
Burdet
Frédéric X
FX
Zaiko
Maïa
M
Hinz
Ursula
U
Zrÿd
Jean-Pierre
JP
D000595:Amino Acid Sequence; D001483:Base Sequence; D050858:Betalains; D020134:Catalytic Domain; D018076:DNA, Complementary / Q000235:genetics; D018744:DNA, Plant / Q000235:genetics; D017343:Genes, Plant; D005816:Genetic Complementation Test; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010105:Oxygenases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny; D010860:Pigments, Biological / Q000096:biosynthesis*; D031822:Portulaca / Q000201:enzymology* / Q000235:genetics; D000644:Quaternary Ammonium Compounds; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D018744:DNA, Plant; D010860:Pigments, Biological; D000644:Quaternary Ammonium Compounds; D050858:Betalains; D010105:Oxygenases; C439587:extradiol dioxygenase