(+)-Maackiain and (+)-medicarpin as phytoalexins in Sophora Japonica and identification of the (−) isomers by biotransformation

VanEtten, Hans D.; Matthews, Patty S.; Mercer, EI

doi:10.1016/s0031-9422(00)80164-8

Cited by 46 publications

(31 citation statements)

References 22 publications

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“…The (-) and (+) enantiomers of maackiain were extracted from roots of red clover and S. japonica, respectively, and purified and crystallized as previously described [19]. (+)Pisatin was extracted from pea [24].…”

Section: Methodsmentioning

confidence: 99%

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Differential toxicity of enantiomers of maackiain and pisatin to phytopathogenic fungi

1992

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Section: Methodsmentioning

confidence: 99%

“…In 19 cases, (+)maackiain was more toxic than (-)maackiain. The isolates of Fusarium reticulatum, which infect a plant that produces both enantiomers [19], were not significantly affected by either compound (Figure 2).…”

Section: Differential Toxicity Of Enantiomersmentioning

confidence: 99%

Differential toxicity of enantiomers of maackiain and pisatin to phytopathogenic fungi

1992

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“…1) (2)(3)(4). This reaction is not identical with the fungal conversion, since the fungus metabolized only (-)maackiain (23) whereas the expected precursor of (+)pisatin, the isomer normally produced by pea, would be the enantiomer (+)maackiain. larity between these two reactions, which appear to play such opposed roles in the interaction between host and parasite, prompted us to examine both of them in this study.…”

mentioning

confidence: 99%

“…and Br. mating population VI isolate T-95, from our culture collection (22,23), was maintained on V-8 juice agar and grown in glucoseasparagine (GA) liquid medium as described (25). Pisum sativum L. cultivar Alaska was grown in a greenhouse.…”

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confidence: 99%

Role of Oxygenases in Pisatin Biosynthesis and in the Fungal Degradation of Maackiain

Matthews¹,

Weiner²,

Matthews³

et al. 1987

Plant Physiol.

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Some isolates of the plant pathogen Nectria haematococca detoxify the isoflavonoid phytoalexin (-)maackiain by hydroxylation at carbon 6a. Precursor feeding studies strongly suggest that the penultimate step in (+)pisatin biosynthesis by Pisum sativum is 6a-hydroxylation of (+)maackiain. We have used "O labeling to test the involvement of oxygenases in these two reactions. When fungal metabolism of maackiain took place under '"02, the product was labeled with 99% efficiency; no label was incorporated by metabolism in H210. Pisatin synthesized by pea pods in the presence of 1802 or H2180 was a mixture of molecules containing up to three labeled oxygen atoms. Primary mass spectra of such mixtures were complex but were greatly simplified by tandem MS. This analysis indicated that the 6a oxygen of pisatin was derived from H20 and not from 02. Labeling patterns for the other five oxygen atoms were consistent with the proposed pathway for biosynthesis of pisatin and related isoflavonoids. We conclude that the fungal hydroxylation of maackiain is catalyzed by an oxygenase, but the biosynthetic route to the 6a hydroxyl of pisatin is unknown.Several pterocarpan phytoalexins are known to be detoxified by Nectria haematococca or by related fungi with the same imperfect state, Fusarium solani. Pisatin is demethylated by a Cyt P-450 monooxygenase (16). Phaseollin is hydroxylated to la-hydroxyphaseollone by a reaction which also appears to be oxygenase-mediated, since 180 labeling has shown that the new oxygen atom is derived from 02 (15). Maackiain and medicarpin are metabolized by one or more of three different initial reactions, depending on the fungal isolate; all three involve the net addition of one oxygen atom (I1, 12). One of these reactions produces the la-hydroxydienone analogous to the phaseollin metabolite. Another, the hydroxylation at carbon 6a, also involves incorporation of an atom from 02, as reported below.Oxygenases are also thought to be responsible for several reactions in the biosynthesis of these phytoalexins. In particular, the 6a-hydroxylation of maackiain has been proposed to be a step in pisatin biosynthesis by Pisum sativum (Fig. 1) (2-4). This reaction is not identical with the fungal conversion, since the fungus metabolized only (-)maackiain (23) whereas the expected precursor of (+)pisatin, the isomer normally produced by pea, would be the enantiomer (+)maackiain. larity between these two reactions, which appear to play such opposed roles in the interaction between host and parasite, prompted us to examine both of them in this study.MATERIALS AND METHODS Biological Materials. Nectria haematococca Berk. and Br. mating population VI isolate T-95, from our culture collection (22, 23), was maintained on V-8 juice agar and grown in glucoseasparagine (GA) liquid medium as described (25). Pisum sativum L. cultivar Alaska was grown in a greenhouse.6a-Hydroxylation of Maackiain. A 3-d culture of N. haematococca in GA medium was filtered, weighed, and suspended in double strength GA medium at 60 m...

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“…In previous work, (+)HMK was obtained by demethylating (+)pisatin using Nectria haematococca (27,28), but yields were low because the fungus further metabolized the (+)HMK. For the current work, (+)HMK was obtained by demethylating (+)pisatin using Aspergillus nidulans III-202, a transformant which contains a gene for pisatin demethylation from N. haematococca T9 (29) and does not metabolize (+)HMK.…”

mentioning

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Purification and Characterization of S-Adenosyl-l-methionine:6a-Hydroxymaackiain 3-O-Methyltransferase from Pisum sativum

Preisig

Matthews²,

VanEtten³

1989

Plant Physiol.

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The isoflavonoid phytoalexin pisatin is synthesized by Pisum sativum in response to microbial infection and certain other forms of stress. An enzyme which synthesizes pisatin by methylating the 3-hydroxyl of (+)6a-hydroxymaackiain (HMK) was extracted from CuCI2-stressed pea seedlings. The enzyme was enriched 370-fold by (NH4)2SO4 precipitation, DEAE chromatography, chromatofocusing, and hydrophobic interaction chromatography (HIC), to a specific activity of 8.2 microkatals per gram protein.Enzyme activity profiles from chromatofocusing and HIC columns suggested the presence of two isozymes, of pi 5.2 and 4.9. Nondenaturing gel filtration of the HIC-purified enzyme gave a single peak of activity at the same elution volume as BSA (66 kilodaltons); the active fractions showed two proteins upon SDS-PAGE, of M, 66,000 and 43,000. The smaller protein was most abundant in chromatographic fractions containing peak enzyme activity throughout purification. In a partially purified preparation, this 43 kilodalton protein was the only one photoaffinity labelled by [3H]S-adenosyl-L-methionine. The purified enzyme preferred the (+) over the (-) stereoisomer of HMK and other pterocarpans; overall, (+)HMK was the best substrate. Km values were 2.3 micromolar for (+)HMK and 35 micromolar for S-adenosyl-L-methionine. The methyltransferase had a pH optimum of 7.9 and no apparent divalent cation requirement.

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(+)-Maackiain and (+)-medicarpin as phytoalexins in Sophora Japonica and identification of the (−) isomers by biotransformation

Cited by 46 publications

References 22 publications

Differential toxicity of enantiomers of maackiain and pisatin to phytopathogenic fungi

Differential toxicity of enantiomers of maackiain and pisatin to phytopathogenic fungi

Role of Oxygenases in Pisatin Biosynthesis and in the Fungal Degradation of Maackiain

Purification and Characterization of S-Adenosyl-l-methionine:6a-Hydroxymaackiain 3-O-Methyltransferase from Pisum sativum

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