Microbial Transformation of ent-Kaurenoic Acid and Its 15-Hydroxy Derivatives by the SG138 Mutant of Gibberella fujikuroi

Abstract

Feeding experiments with ent-kaurenoic acid (4), 15R-hydroxy-ent-kaurenoic acid (5), 15 -hydroxy-entkaurenoic acid (6), and mixtures of 4 plus 5 and 4 plus 6 were conducted using the SG138 mutant of Gibberella fujikuroi, to gain information about the phenotype of this unique strain. The biotransformation of 5 gave 7 ,15R-dihydroxykaurenolide (9) and 7 ,15R-dihydroxy-ent-kaurenoic acid (13). The incubation of 6 produced 7 ,15 -dihydroxy-ent-kaurenoic acid (7) and 7 ,15 -dihydroxykaurenolide (10). No 15-hydroxylated gibberellins were detected in any of these experiments. The results indicated that a hydroxy group at C-15 does not inhibit 7 -hydroxylase activity but in the SG138 strain obstructs the enzymatic ring-B contraction of ent-kaurenoids to gibberellins. Exogenous 4 stimulated both the excretion of ent-kaurene and the fungal metabolism of 5 and 6. Gibberellins (GAs) are terpenoid phytohormones which play an important role in the regulation of plant growth and development. Apart from plants some fungi also produce these terpenoids, particularly Gibberella fujikuroi (Hypocreale). GAs have many applications in agriculture and the brewing industry. 1 Therefore, their abundant production by some strains of G. fujikuroi has led to the study of their biosynthesis by this fungus in considerable detail over the last three decades. 2 Some aspects of the biogenesis of the gibberellins via ent-kaurene (1), entkaurenol (2), ent-kaurenal (3), and ent-kaurenoic acid (4) are fully described, 2 but many of the biochemical and physiological aspects of fungal gibberellin biosynthesis are still poorly understood. As part of our research into the biotechnology of G. fujikuroi we have investigated the metabolites produced by the GA-producer IMI58289 strain. 3,4 Subsequently, the kaurenoids and GAs synthesized by several gib-mutants developed from the IMI58289 wildtype strain were also analyzed. 5 The results of these analyses and preliminary feeding experiments 6 suggested to us that one of these mutants (SG138) was blocked in the oxidative transformations from ent-kaurene (1) to entkaurenoic acid (4), as well as in 3 -hydroxylation, 13 -hydroxylation, and the loss of C-20 by the GAs. These unusual features prompted us to make further feeding experiments with strain SG138 to gain more information about its unusual phenotype. We describe here the results of incubations with ent-kaurenoic acid (4), the "unnatural" substrates 15R-hydroxy-ent-kaurenoic acid (5) and 15 -hydroxy-ent-kaurenoic acid (6), and mixtures of 4 plus 5 and 4 plus 6. Throughout these experiments it was observed that ent-kaurenoic acid (4) substantially stimulated the metabolism and/or excretion of 1, 5, and 6.The results of incubations using the SG138 mutant of G. fujikuroi, both with and without ent-kaurenoic acid (4), are summarized in Table 1. All compounds included in this table were identified by means of GC-MS. Several substances could be isolated, and their structures were confirmed by NMR techniques (see the Experimental Section for more details). A num...

The NADPH-cytochrome P450 Reductase Gene from Gibberella fujikuroi Is Essential for Gibberellin Biosynthesis

“…The GA-deficient mutant G. fujikuroi SG138 (kindly provided by J. Avalos, University of Sevilla, Spain) strain was derived from IMI58289 via UV mutagenesis of spores (37).…”

“…Thus, loss of CPR activity should affect the rates of several reactions in the pathway. Such an effect was found for the UV-induced G. fujikuroi mutant SG138, which has lost most of the oxidation steps catalyzed by P450 monooxygenases (37). Because it is unlikely that all structural P450 monooxygenase genes are mutated by one UV treatment it is possible that the mutant contains a lesion mutation in the cpr gene.…”

Smallanthus sonchifolius and Lepidium meyenii - prospective Andean crops for the prevention of chronic diseases

“…Its antifungal activity has also been attributed to 4-hydroxystyrene and 3,4-dihydroxystyrene that are formed in yacon damaged leaves by oxidative decarboxylation of p-coumaric and caffeic acids by enzymatic systems of epiphytic bacteria 52,53 . Ent-kaurenoic acid is one of the terpenoid phytohormone gibberellins biosynthesis intermediates in Gibberella fujikuroi 54 . Entkaurene is responsible for the antibacterial activity of Brazilian propolis from native stingless bees Melipona quadrifasciata anthidioides 55 .…”

New stereoisomeric derivatives of jasmonic acid generated by biotransformation with the fungus Gibberella fujikuroi affect the viability of human cancer cells

“…One the best characterized biotransformation systems using G. fujikuroi is the chemical modification of some precursors of the plant hormone gibberellins (Barrero et al 2001;Fraga et al 2004;Fraga et al 2007). The fungus shows the presence of enzymes involved in the biosynthetic pathway of such type of diterpenoids which might be responsible for the chemical modification of the gibberellin structure (Tudzynski et al 2003;Malonek et al 2004;Tudzynski, 2005).…”