131887-80-4Relevant academic research and scientific papers
Catalytic specificity of pea O-methyltransferases suggests gene duplication for (+)-pisatin biosynthesis
Akashi, Tomoyoshi,VanEtten, Hans D.,Sawada, Yuji,Wasmann, Catherine C.,Uchiyama, Hiroshi,Ayabe, Shin-ichi
, p. 2525 - 2530 (2007/10/03)
S-adenosyl-l-methionine: 2-hydroxyisoflavanone 4′-O-methyltransferase (HI4′OMT) methylates 2,7, 4′-trihydroxyisoflavanone to produce formononetin, an essential intermediate in the synthesis of isoflavonoids with methoxy or methylenedioxy groups at carbon 4′ (isoflavone numbering). HI4′OMT is highly similar (83% amino acid identity) to (+)-6a-hydroxymaackiain 3-O-methyltransferase (HMM), which catalyzes the last step of (+)-pisatin biosynthesis in pea. Pea contains two linked copies of HMM with 96% amino acid identity. In this report, the catalytic activities of the licorice HI4′OMT protein and of extracts of Escherichia coli containing the pea HMM1 or HMM2 protein are compared on 2,7,4′-trihydroxyisoflavanone and enantiomers of 6a-hydroxymaackiain. All these enzymes produced radiolabelled 2,7-dihydroxy-4′-methoxyisoflavanone or (+)-pisatin from 2,7,4′-trihydroxyisoflavanone or (+)-6a-hydroxymaakiain when incubated with [methyl-14C]-S-adenosyl-l-methionine. No product was detected when (-)-6a-hydroxymaackiain was used as the substrate. HI4′OMT and HMM1 showed efficiencies (relative Vmax/Km) for the methylation of 2,7,4′-trihydroxyisoflavanone 20 and 4 times higher than for the methylation of (+)-6a-hydroxymaackiain, respectively. In contrast, HMM2 had a higher Vmax and lower Km on (+)-6a-hydroxymaackiain, and had a 67-fold higher efficiency for the methylation of (+)-6a-hydroxymaackiain than that for 2,7,4′-trihydroxyisoflavanone. Among the 15 sites at which HMM1 and HMM2 have different amino acid residues, 11 of the residues in HMM1 are the same as found in HI4′OMTs from three plant species. Modeling of the HMM proteins identified three or four putative active site residues responsible for their different substrate preferences. It is proposed that HMM1 is the pea HI4′OMT and that HMM2 evolved by the duplication of a gene encoding a general biosynthetic enzyme (HI4′OMT).
P-450-DEPENDENT OXIDATIVE REARRANGEMENT IN ISOFLAVONE BIOSYNTHESIS: RECONSTITUTION OF P-450 AND NADPH:P-450 REDUCTASE
Hakamatsuka, Takashi,Hashim, Muhammed Faisal,Ebizuka, Yutaka,Sankawa, Ushio
, p. 5969 - 5978 (2007/10/02)
The reaction mechanism of oxidative rearrangement in the conversion of liquiritigenin, a flavanone, into 2,7,4'-trihydroxyisoflavanone was studied in elicitor-challenged Pueraria lobata cell cultures.The involvement of cytochrome P-450 in the reaction, hydroxylation associated with 1,2-aryl migration, was proved previously by the inhibition experiments with carbon monoxide and P-450 inhibitors.In order to obtain rigorous evidence proving that the enzyme is a P-450, a reconstitution experiment was performed with solubilized cytochrome P-450 and NADPH:cytochrome P-450 reductase fractions.During these studies we noticed that various biosynthetic reactions can be interpreted as P-450-mediated reactions associated with migration or bond cleavage.Ring contraction of 7-hydroxy-kaurenoic acid in gibberellin biosynthesis, the formation of a furan ring in furanocoumarin biosynthesis and several rearrangement reactions in steroid metabolism are discussed as examples of P-450 reactions associated with migration or bond cleavage.
