Characterization of the last step of the aerobic phenylacetic acid degradation pathway

Nogales, Juan; Macchi, Raffaella; Franchi, Federico; Barzaghi, D; Fernández, Cristina; Garcı́a, José Luis; Bertoni, Giovanni; Dı́az, Eduardo

doi:10.1099/mic.0.2006/002444-0

Cited by 57 publications

(28 citation statements)

References 44 publications

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“…The observation that growth of the deletion strains on the aromatic quinate was completely abolished is of particular interest. This result was surprising because quinate is metabolized to acetyl-CoA and succinyl-CoA via the protocatechuate pathway (as characterized in bacteria [32]), which has been analyzed by the isolation of pca mutants unable to grow on either quinate or benzoate (25). The genes for the enzymes of this pathway have not been cloned.…”

Section: Resultsmentioning

confidence: 99%

ATP-Citrate Lyase Is Required for Production of Cytosolic Acetyl Coenzyme A and Development in Aspergillus nidulans

Hynes

Murray

2010

Eukaryot Cell

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Acetyl coenzyme A (CoA) is a central metabolite in carbon and energy metabolism and in the biosynthesis of cellular molecules. A source of cytoplasmic acetyl-CoA is essential for the production of fatty acids and sterols and for protein acetylation, including histone acetylation in the nucleus. In Saccharomyces cerevisiae and Candida albicans acetyl-CoA is produced from acetate by cytoplasmic acetyl-CoA synthetase, while in plants and animals acetyl-CoA is derived from citrate via ATP-citrate lyase. In the filamentous ascomycete Aspergillus nidulans, tandem divergently transcribed genes (aclA and aclB) encode the subunits of ATP-citrate lyase, and we have deleted these genes. Growth is greatly diminished on carbon sources that do not result in cytoplasmic acetyl-CoA, such as glucose and proline, while growth is not affected on carbon sources that result in the production of cytoplasmic acetyl-CoA, such as acetate and ethanol. Addition of acetate restores growth on glucose or proline, and this is dependent on facA, which encodes cytoplasmic acetyl-CoA synthetase, but not on the regulatory gene facB. Transcription of aclA and aclB is repressed by growth on acetate or ethanol. Loss of ATP-citrate lyase results in severe developmental effects, with the production of asexual spores (conidia) being greatly reduced and a complete absence of sexual development. This is in contrast to Sordaria macrospora, in which fruiting body formation is initiated but maturation is defective in an ATP-citrate lyase mutant. Addition of acetate does not repair these defects, indicating a specific requirement for high levels of cytoplasmic acetyl-CoA during differentiation. Complementation in heterokaryons between aclA and aclB deletions for all phenotypes indicates that the tandem gene arrangement is not essential.

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Section: Resultsmentioning

confidence: 99%

ATP-Citrate Lyase Is Required for Production of Cytosolic Acetyl Coenzyme A and Development in Aspergillus nidulans

Hynes

Murray

2010

Eukaryot Cell

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“…Many attempts to identify pathway intermediates other than phenylacetyl-CoA led only to rough outlines (14,(18)(19)(20). We set out to elucidate the pathway for phenylacetate degradation by producing purified enzymes coded by the E. coli K12 and Pseudomonas sp.…”

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confidence: 99%

Bacterial phenylalanine and phenylacetate catabolic pathway revealed

Teufel

Mascaraque²,

Ismail³

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

313

339

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Aromatic compounds constitute the second most abundant class of organic substrates and environmental pollutants, a substantial part of which (e.g., phenylalanine or styrene) is metabolized by bacteria via phenylacetate. Surprisingly, the bacterial catabolism of phenylalanine and phenylacetate remained an unsolved problem. Although a phenylacetate metabolic gene cluster had been identified, the underlying biochemistry remained largely unknown. Here we elucidate the catabolic pathway functioning in 16% of all bacteria whose genome has been sequenced, including Escherichia coli and Pseudomonas putida. This strategy is exceptional in several aspects. Intermediates are processed as CoA thioesters, and the aromatic ring of phenylacetyl-CoA becomes activated to a ring 1,2-epoxide by a distinct multicomponent oxygenase. The reactive nonaromatic epoxide is isomerized to a seven-member O-heterocyclic enol ether, an oxepin. This isomerization is followed by hydrolytic ring cleavage and β-oxidation steps, leading to acetyl-CoA and succinyl-CoA. This widespread paradigm differs significantly from the established chemistry of aerobic aromatic catabolism, thus widening our view of how organisms exploit such inert substrates. It provides insight into the natural remediation of man-made environmental contaminants such as styrene. Furthermore, this pathway occurs in various pathogens, where its reactive early intermediates may contribute to virulence.enoyl-CoA hydratase | epoxide | oxepin | oxygenase | phenylacetic acid T he biggest challenge for organisms using aromatic compounds as growth substrates is to overcome the stabilizing resonance energy of the aromatic ring system. This aromatic structure makes the substrates unreactive toward oxidation or reduction and thus requires elaborate degradation strategies. How microorganisms cope with this problem depends primarily on the availability of oxygen (1). Aerobic pathways use oxygen both for hydroxylation and for cleavage of the ring (2, 3). In contrast, under anaerobic conditions the common strategy consists of activation by CoA-thioester formation, shortening of the side chain, and energy-driven ring reduction, which also applies to phenylacetate catabolism (ref. 4 and literature cited therein).The aerobic strategy is illustrated by the metabolism of phenylacetate and phenylacetyl-CoA, which are derived from a variety of substrates such as phenylalanine, lignin-related phenylpropane units, 2-phenylethylamine, phenylalkanoic acids with an even number of carbon atoms, or even environmental contaminants such as styrene and ethylbenzene (5-7). Rarely, phenylalanine is hydroxylated to tyrosine, which can be converted into 4-hydroxyphenylpyruvate, followed by hydroxylation to homogentisate (2,5-dihydroxyphenylacetate) as the central intermediate. The aromatic ring of homogentisate then is split by a ring-cleaving homogentisate dioxygenase, and finally fumarate and acetoacetate are produced (8). In most cases, however, phenylalanine is converted into phenylacetate. A conventional aero...

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“…The PA-activating enzyme, phenylacetyl-coenzyme A (PA-CoA) ligase, PaaK (15), and the PA-CoA ring hydroxylation system, comprised of PaaA, PaaB, PaaC, PaaD, and PaaE (15,26), are involved in the first and second steps, respectively. The third step, the opening of the aromatic ring, may be performed by PaaZ (15) or by PaaZ, PaaG, and PaaJ (26), followed by further degradation of the resulting aliphatic compound through a ␤-oxidation-like pathway complex by PaaF, PaaH (26), and PaaJ (41).…”

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confidence: 99%

A Functional Phenylacetic Acid Catabolic Pathway Is Required for Full Pathogenicity of Burkholderia cenocepacia in the Caenorhabditis elegans Host Model

et al. 2008

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Burkholderia cenocepacia is a member of the Burkholderia cepacia complex, a group of metabolically versatile bacteria that have emerged as opportunistic pathogens in cystic fibrosis and immunocompromised patients. Previously a screen of transposon mutants in a rat pulmonary infection model identified an attenuated mutant with an insertion in paaE, a gene related to the phenylacetic acid (PA) catabolic pathway. In this study, we characterized gene clusters involved in the PA degradation pathway of B. cenocepacia K56-2 in relation to its pathogenicity in the Caenorhabditis elegans model of infection. We demonstrated that targeted-insertion mutagenesis of paaA and paaE, which encode part of the putative PA-coenzyme A (CoA) ring hydroxylation system, paaZ, coding for a putative ring opening enzyme, and paaF, encoding part of the putative beta-oxidation system, severely reduces growth on PA as a sole carbon source. paaA and paaE insertional mutants were attenuated for virulence, and expression of paaE in trans restored pathogenicity of the paaE mutant to wild-type levels. Interruption of paaZ and paaF slightly increased virulence. Using gene interference by ingested doublestranded RNA, we showed that the attenuated phenotype of the paaA and paaE mutants is dependent on a functional p38 mitogen-activated protein kinase pathway in C. elegans. Taken together, our results demonstrate that B. cenocepacia possesses a functional PA degradation pathway and that the putative PA-CoA ring hydroxylation system is required for full pathogenicity in C. elegans.

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Characterization of the last step of the aerobic phenylacetic acid degradation pathway

Cited by 57 publications

References 44 publications

ATP-Citrate Lyase Is Required for Production of Cytosolic Acetyl Coenzyme A and Development in Aspergillus nidulans

ATP-Citrate Lyase Is Required for Production of Cytosolic Acetyl Coenzyme A and Development in Aspergillus nidulans

Bacterial phenylalanine and phenylacetate catabolic pathway revealed

A Functional Phenylacetic Acid Catabolic Pathway Is Required for Full Pathogenicity of Burkholderia cenocepacia in the Caenorhabditis elegans Host Model

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