Hydroxylation of the native brassinosteroids 24-epicastasterone and 24-epibrassinolide by the fungus Cunninghamella echinulata

Voigt, B.; Porzel, Andrea; Naumann, Heidrun; Hörhold‐Schubert, Cläre; Adam, Günter

doi:10.1016/0039-128x(93)90091-z

Cited by 20 publications

(9 citation statements)

References 11 publications

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“…Although steroids are common exogenous substrates for filamentous fungi, hydroxylated at the 11␣-, 11␤-, and 16␣-positions in most cases, 12␤-hydroxylation is relatively rare (18). Examples include hydroxylation of 24-epi-brassinolide and 24-epi-castasterone by Cunninghamella, of pregn-4-ene-3,20-dione by Cephalosporium, of mexrenone by Mortierella isabellina, and of scillarenin by Rhizopus arrhizus (6,7,21,25). These reactions are not specific, however, and are coupled with hydroxylation at other sites.…”

Section: Discussionmentioning

confidence: 99%

Specific 12β-Hydroxylation of Cinobufagin by Filamentous Fungi

Guo

et al. 2004

Appl Environ Microbiol

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Biotransformation of natural products has great potential for producing new drugs and could provide in vitro models of mammalian metabolism. Microbial transformation of the cytotoxic steroid cinobufagin was investigated. Cinobufagin could be specifically hydroxylated at the 12␤-position by the fungus Alternaria alternata. Six products from a scaled-up fermentation were obtained by silica gel column chromatography and reversed-phase liquid chromatography and were identified as 12␤-hydroxyl cinobufagin, 12␤-hydroxyl desacetylcinobufagin, 3-oxo-12␤-hydroxyl cinobufagin, 3-oxo-12␤-hydroxyl desacetylcinobufagin, 12-oxo-cinobufagin, and 3-oxo-12␣-hydroxyl cinobufagin. The last five products are new compounds. 12␤-Hydroxylation of cinobufagin by A. alternata is a fast catalytic reaction and was complete within 8 h of growth with the substrate. This reaction was followed by dehydrogenation of the 3-hydroxyl group and then deacetylation at C-16. Hydroxylation at C-12␤ also was the first step in the metabolism of cinobufagin by a variety of fungal strains. In vitro cytotoxicity assays suggest that 12␤-hydroxyl cinobufagin and 3-oxo-12␣-hydroxyl cinobufagin exhibit somewhat decreased but still significant cytotoxic activities. The 12␤-hydroxylated bufadienolides produced by microbial transformation are difficult to obtain by chemical synthesis.Bufadienolides are steroids with a characteristic ␣-pyrone ring at the C-17 position; they have cardiotonic, blood pressure-stimulating, antiviral, and local anesthetic activities (20). More than 300 bufadienolides have been isolated from natural sources including plants and animals (15,23). These compounds have been reported to have significant antitumor activities (9,19,31,33). Cinobufagin (compound I) is a bufadienolide with a 14␤,15␤-epoxy ring, originally isolated as a major component of the traditional Chinese drug, Chan'Su (also called toad venom or toad poison), which is prepared from the skin secretions of giant toads (10). Cinobufagin can induce apoptosis and elevate intracellular Ca 2ϩ levels (13,14). Against cancer cells it has a 50% inhibitory concentration (IC 50 ) of approximately 10 Ϫ8 mol/liter (30). Unfortunately, it is poorly soluble in water and is toxic to humans; therefore, analogs with improved pharmaceutical properties, such as those obtained from plant cell suspension cultures (27,29), are needed before this compound can be utilized effectively in a clinical setting.Microbial transformation is an important tool for structural modification of organic compounds, especially natural products with complicated structures (17,22). It can be used to synthesize chemical structures that are difficult to obtain by other means (24) and as a model of mammalian metabolism due to the similarity between mammalian and microbial cytochrome P 450 enzyme systems (1,2,4,5,12). The metabolism of cinobufagin in rat liver microsomes produced at least six metabolites (32), resulting primarily from deacetylation at C-16 and epimerization of 3-OH via a 3-keto intermediate. The exact s...

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Section: Discussionmentioning

confidence: 99%

Specific 12β-Hydroxylation of Cinobufagin by Filamentous Fungi

Guo

et al. 2004

Appl Environ Microbiol

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“…Diglycosidation of exogenous 24-epiteasterone occurred in cell cultures of tomato, Lycopersicon esculentum (60), and in the same system, hydroxylation at the 25-and 26-positions of 24-epibrassinolide required two separate hydroxylases, with only the 25-hydroxylase sensitive to cytochrome P450 inhibitors (127). How many of these transformations occur in vivo as part of normal biosynthesis and turnover of BRs remains to be seen, but the possibility of very low levels of even more active members of the family, like those produced by fungal metabolism (48,117,118) is tantalizing.…”

Section: Conjugation and Metabolismmentioning

confidence: 95%

BRASSINOSTEROIDS: Essential Regulators of Plant Growth and Development

Clouse

Sasse

1998

Annu. Rev. Plant. Physiol. Plant. Mol. Biol.

1,266

776

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Brassinosteroids (BRs) are growth-promoting natural products found at low levels in pollen, seeds, and young vegetative tissues throughout the plant kingdom. Detailed studies of BR biosynthesis and metabolism, coupled with the recent identification of BR-insensitive and BR-deficient mutants, has greatly expanded our view of steroids as signals controlling plant growth and development. This review examines the microchemical and molecular genetic analyses that have provided convincing evidence for an essential role of BRs in diverse developmental programs, including cell expansion, vascular differentiation, etiolation, and reproductive development. Recent advances relevant to the molecular mechanisms of BR-regulated gene expression and BR signal transduction are also discussed.

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“…include b-epimerization of 2a-OH and 3a-OH (Fujioka, 1999;Nishikawa et al, 1995;Suzuki et al, 1995), hydroxylation of C-12, C-20, C-25 and C-26 (Hai et al, 1996;Kolbe et al, 1996;Neff et al, 1999;Voigt et al, 1993;Winter et al, 1997;Yokota et al, 1996), demethylation of C-26 CH 3 and C-28 CH 3 (Fujioka et al, 2000b;Kim et al, 2000;Watanabe et al, 2001), side chain cleavage of C-20/22 (Kolbe et al, 1996), oxidation of 23-OH , sulfonation of 22-OH (Rouleau et al, 1999), conjugation with fatty acids at 3b-OH (Asakawa et al, 1996;Kolbe et al, 1995), conjugation with glucose at 2a-OH, 3b-OH, 23-OH, 25-OH and 26-OH (Fujioka, 1999;Hai et al, 1996;Kolbe et al, 1998;Soeno et al, 2000;Suzuki et al, 1993;Winter et al, 1997), and conjugation with 6-O-b glucosylglucose or 4-O-b glucosylgalactose at 3b-OH (Kolbe et al, 1998). Only three enzymes have been identified to be responsible for BR-specific inactivation reactions: (1) a steroid sulfotransferase from Brassica napus that is responsible for sulfonation at 22-OH of 24-epicathasterone (Rouleau et al, 1999), (2) an Arabidopsis BAS1 that participates in a hydroxylation reaction at C-26 (Neff et al, 1999;Turk et al, 2003), and (3) another Arabidopsis protein UGT73C5 that catalyzes the conjugation of BL with glucose at 23-OH (Poppenberger et al, 2005).…”

Section: ) Reactions Putatively Involved In Br Inactivationmentioning

confidence: 99%

BEN1, a gene encoding a dihydroflavonol 4‐reductase (DFR)‐like protein, regulates the levels of brassinosteroids in Arabidopsis thaliana

et al. 2007

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SummaryThe ben1-1D (bri1-5 enhanced 1-1dominant) mutant was identified via an activation-tagging screen for bri1-5 extragenic modifiers. bri1-5 is a weak mutant allele of the brassinosteroid receptor gene, BRI1. Overexpression of BEN1 greatly enhances the defective phenotypes of bri1-5 plants. Removal of BEN1 by gene disruption in a Col-0 wild-type background, on the other hand, promotes the elongation of organs. Because BEN1 encodes a novel protein homologous to dihydroflavonol 4-reductase (DFR) and anthocyanidin reductase (BAN), BEN1 is probably involved in a brassinosteroid metabolic pathway. Analyses of brassinosteroid profiles demonstrated that BEN1 is indeed responsible for regulating the levels of several brassinosteroids, including typhasterol, castasterone and brassinolide. In vivo feeding and in vitro biochemical assays suggest that BEN1 is probably involved in a new mechanism to regulate brassinosteroid levels. These results provide additional insight into the regulatory mechanisms of bioactive brassinosteroids.

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Hydroxylation of the native brassinosteroids 24-epicastasterone and 24-epibrassinolide by the fungus Cunninghamella echinulata

Cited by 20 publications

References 11 publications

Specific 12β-Hydroxylation of Cinobufagin by Filamentous Fungi

Specific 12β-Hydroxylation of Cinobufagin by Filamentous Fungi

BRASSINOSTEROIDS: Essential Regulators of Plant Growth and Development

BEN1, a gene encoding a dihydroflavonol 4‐reductase (DFR)‐like protein, regulates the levels of brassinosteroids in Arabidopsis thaliana

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