Novel Scheme for Biosynthesis of Aryl Metabolites from l -Phenylalanine in the Fungus Bjerkandera adusta

Abstract

Aryl metabolite biosynthesis was studied in the white rot fungus Bjerkandera adusta cultivated in a liquid medium supplemented with L-phenylalanine. Aromatic compounds were analyzed by gas chromatographymass spectrometry following addition of labelled precursors ( 14 C-and 13 C-labelled L-phenylalanine), which did not interfere with fungal metabolism. The major aromatic compounds identified were benzyl alcohol, benzaldehyde (bitter almond aroma), and benzoic acid. Hydroxy-and methoxybenzylic compounds (alcohols, aldehydes, and acids) were also found in fungal cultures. Intracellular enzymatic activities (phenylalanine ammonia lyase, aryl-alcohol oxidase, aryl-alcohol dehydrogenase, aryl-aldehyde dehydrogenase, lignin peroxidase) and extracellular enzymatic activities (aryl-alcohol oxidase, lignin peroxidase), as well as aromatic compounds, were detected in B. adusta cultures. Metabolite formation required de novo protein biosynthesis. Our results show that L-phenylalanine was deaminated to trans-cinnamic acid by a phenylalanine ammonia lyase and trans-cinnamic acid was in turn converted to aromatic acids (phenylpyruvic, phenylacetic, mandelic, and benzoylformic acids); benzaldehyde was a metabolic intermediate. These acids were transformed into benzaldehyde, benzyl alcohol, and benzoic acid. Our findings support the hypothesis that all of these compounds are intermediates in the biosynthetic pathway from L-phenylalanine to aryl metabolites. Additionally, transcinnamic acid can also be transformed via ␤-oxidation to benzoic acid. This was confirmed by the presence of acetophenone as a ␤-oxidation degradation intermediate. To our knowledge, this is the first time that a ␤-oxidation sequence leading to benzoic acid synthesis has been found in a white rot fungus. A novel metabolic scheme for biosynthesis of aryl metabolites from L-phenylalanine is proposed.Consumer preferences for products with a natural origin have led to the exploitation of microbial sources that produce natural aroma compounds (16,28). Among the potential aroma producers, white rot basidiomycetes are probably the most versatile microorganisms. These fungi are able to produce a wide variety of volatile aryl metabolites of commercial interest, such as vanillin, benzaldehyde (bitter almond aroma), and cinnamaldehyde (1,7,12). Therefore, fermentation of natural substrates, such as L-phenylalanine or tyrosine, by white rot fungi can offer alternative routes for biosynthesis of a wide spectrum of aryl metabolites (11, 21). Other biosynthetic precursors, like aromatic acids, stimulate the production of aryl metabolites in Bjerkandera adusta BOS55 (21).The metabolism of L-phenylalanine has been studied in several white rot fungi (13, 17). Among the extracellular aromatic compounds that these organisms produce, veratryl alcohol has received special attention because it is known to be a substrate and possibly a mediator in lignin biodegradation (6,15). This compound is the major aryl metabolite formed in Phanerochaete chrysosporium cultures supplemented wi...

Role of the Phenylalanine-Hydroxylating System in Aromatic Substance Degradation and Lipid Metabolism in the Oleaginous Fungus Mortierella alpina

“…M. alpina was unable to grow on medium containing phenylacetate as the sole carbon source (see Fig. S1C), even if it was incubated for over 2 weeks or at a higher phenylacetate concentration (1 M), which is in contrast to other fungi, such as P. chrysogenum, A. nidulans, and A. niger (17)(18)(19)(20). Thus, these results show that M. alpina can degrade phenylalanine and tyrosine, but not phenylacetate.…”

“…The conventional aerobic route for phenylacetate, which leads to homogentisate, occurs in fungi (Fig. 2) (17)(18)(19)(20). However, the M. alpina genome sequence lacks an ortholog of phenylacetate 2-hydroxylase (EC 1.14.13.-) to catalyze the hydroxylation of phenylacetate to homogentisate (Fig.…”

“…2A) or into cinnamic acid in Bjerkandera adusta and Schizophyllum commune ( Fig. 2C) (17)(18)(19)(20). The cinnamate thus formed is converted to protocatechuate through benzoate, whereas phenylpyruvate is converted to homogentisate, which is catabolized by cleavage of the aromatic ring to yield fumarate and acetyl-CoA.…”

Bioconversion of cinnamic acid derivatives by Schizophyllum commune

“…However, the fungus hydrogenated the double bond fi rst in p-coumaric acid (Fig. 5B).Reduction of p-coumaric and ferulic acids by whiterot fungi has been reported previously; for example, the reduction of the carboxyl group at the γ-position (Enoki et al, 1981;Estrada Alvarado et al, 2001Falconnier et al, 1994;Gupta et al, 1981;Krings et al, 2001;Lapadatescu et al, 2000;Nishida and Fukuzumi, 1978) and the hydrogenation of C α -C β double bond (Enoki et al, 1981;Estrada Alvarado et al, 2001;Gupta et al, 1981). This study demonstrates that S. commune reduced the side chain of p-coumaric acid but not that of ferulic acid ( Fig.…”

“…Modifi cation of the side chain of cinnamic acid by other white-rot Basidiomycetes has been reported. Reduction of cinnamic acid into cinnamyl aldehyde was determined during L-phenylalanine metabolism by Bjerkandera adusta (Lapadatescu et al, 2000). Formation of benzaldehyde and benzoic acid from cinnamic acid has been reported in Phanerochaete chrysosporium and B. adusta (Jensen et al, 1994;Lapadatescu et al, 2000).…”