Pathways for degradation of lignin in bacteria and fungi

Summary

Analysis of the catabolic potential of Pseudomonas putida KT2440 against a wide range of natural aromatic compounds and sequence comparisons with the entire genome of this microorganism predicted the existence of at least four main pathways for the catabolism of central aromatic intermediates, that is, the protocatechuate (pca genes) and catechol (cat genes) branches of the β-ketoadipate pathway, the homogentisate pathway (hmg/fah/mai genes) and the phenylacetate pathway (pha genes). Two additional gene clusters that might be involved in the catabolism of N-heterocyclic aromatic compounds (nic cluster) and in a central meta-cleavage pathway (pcm genes) were also identified. Furthermore, the genes encoding the peripheral pathways for the catabolism of p-hydroxybenzoate (pob), benzoate (ben), quinate (qui), phenylpropenoid compounds (fcs, ech, vdh, cal, van, acd and acs), phenylalanine and tyrosine (phh, hpd) and n-phenylalkanoic acids (fad) were mapped in the chromosome of P. putida KT2440. Although a repetitive extragenic palindromic (REP) element is usually associated with the gene clusters, a supraoperonic clustering of catabolic genes that channel different aromatic compounds into a common central pathway (catabolic island) was not observed in P. putida KT2440. The global view on the mineralization of aromatic compounds by P. putida KT2440 will facilitate the rational manipulation of this strain for improving biodegradation/biotransformation processes, and reveals this bacterium as a useful model system for studying biochemical, genetic, evolutionary and ecological aspects of the catabolism of aromatic compounds.

Genomic analysis of the aromatic catabolic pathways from Pseudomonas putida KT2440

Lignins are the most abundant aromatic compounds in nature, and their decomposition is essential to the terrestrial carbon cycle. White rot fungi secreting phenol oxidases are assumed to be involved in the initial degradation of native lignin, whereas bacteria play a main role in the mineralization of lignin-derived low-molecular-weight compounds in soil. There are a number of reports on the degradation pathways for lignin-derived aromatic compounds, but their catabolism has not been enzymatically or genetically characterized. Sphingomonas paucimobilis SYK-6 is one of the best-characterized lignin-degrading bacteria. It can grow on a wide variety of lignin-related biaryls and monoaryls, including beta-aryl ether, biphenyl, diarylpropane, and phenylpropane. These compounds are degraded via the protocatechuate (PCA) 4,5-cleavage pathway or multiple 3-O-methylgallate (3MGA) catabolic pathways. In this review, the enzyme systems for beta-aryl ether and biphenyl degradation, O demethylation linked with one carbon metabolism, the PCA 4,5-cleavage pathway, and the multiple 3MGA catabolic pathways in SYK-6 are outlined.

https://www.tandfonline.com/doi/pdf/10.1271/bbb.60437

Genetic and Biochemical Investigations on Bacterial Catabolic Pathways for Lignin-Derived Aromatic Compounds

Vanillin is one of the world's most important flavor compounds, with a global market of 180 million dollars. Natural vanillin is derived from the cured seed pods of the vanilla orchid (Vanilla planifolia), but most of the world's vanillin is synthesized from petrochemicals or wood pulp lignins. We have established a true de novo biosynthetic pathway for vanillin production from glucose in Schizosaccharomyces pombe, also known as fission yeast or African beer yeast, as well as in baker's yeast, Saccharomyces cerevisiae. Productivities were 65 and 45 mg/liter, after introduction of three and four heterologous genes, respectively. The engineered pathways involve incorporation of 3-dehydroshikimate dehydratase from the dung mold Podospora pauciseta, an aromatic carboxylic acid reductase (ACAR) from a bacterium of the Nocardia genus, and an O-methyltransferase from Homo sapiens. In S. cerevisiae, the ACAR enzyme required activation by phosphopantetheinylation, and this was achieved by coexpression of a Corynebacterium glutamicum phosphopantetheinyl transferase. Prevention of reduction of vanillin to vanillyl alcohol was achieved by knockout of the host alcohol dehydrogenase ADH6. In S. pombe, the biosynthesis was further improved by introduction of an Arabidopsis thaliana family 1 UDP-glycosyltransferase, converting vanillin into vanillin beta-D-glucoside, which is not toxic to the yeast cells and thus may be accumulated in larger amounts. These de novo pathways represent the first examples of one-cell microbial generation of these valuable compounds from glucose. S. pombe yeast has not previously been metabolically engineered to produce any valuable, industrially scalable, white biotech commodity.

De novo biosynthesis of vanillin in fission yeast (Schizosaccharomyces pombe) and baker's yeast (Saccharomyces cerevisiae).

Although Escherichia coli has long been recognized as the best-understood living organism, little was known about its abilities to use aromatic compounds as sole carbon and energy sources. This review gives an extensive overview of the current knowledge of the catabolism of aromatic compounds by E. coli. After giving a general overview of the aromatic compounds that E. coli strains encounter and mineralize in the different habitats that they colonize, we provide an up-to-date status report on the genes and proteins involved in the catabolism of such compounds, namely, several aromatic acids (phenylacetic acid, 3- and 4-hydroxyphenylacetic acid, phenylpropionic acid, 3-hydroxyphenylpropionic acid, and 3-hydroxycinnamic acid) and amines (phenylethylamine, tyramine, and dopamine). Other enzymatic activities acting on aromatic compounds in E. coli are also reviewed and evaluated. The review also reflects the present impact of genomic research and how the analysis of the whole E. coli genome reveals novel aromatic catabolic functions. Moreover, evolutionary considerations derived from sequence comparisons between the aromatic catabolic clusters of E. coli and homologous clusters from an increasing number of bacteria are also discussed. The recent progress in the understanding of the fundamentals that govern the degradation of aromatic compounds in E. coli makes this bacterium a very useful model system to decipher biochemical, genetic, evolutionary, and ecological aspects of the catabolism of such compounds. In the last part of the review, we discuss strategies and concepts to metabolically engineer E. coli to suit specific needs for biodegradation and biotransformation of aromatics and we provide several examples based on selected studies. Finally, conclusions derived from this review may serve as a lead for future research and applications.

http://envismadrasuniv.org/Bioremediation/pdf/Biodegradation%20of%20aromatic.pdf

Biodegradation of Aromatic Compounds by Escherichia coli

Transposon Tn5 genomic mutants of plant-growth-promoting Pseudomonas putida strain WCS358 have been isolated which no longer utilize ferulic and coumaric acids as sole sources of carbon and energy. Genetic studies confirmed previous biochemical data showing that ferulic acid is degraded via vanillic acid, and coumaric acid via hydroxybenzoic acid. The genes involved in these enzymic steps were cloned and characterized. Two proteins designated Fca (26.5 kDa) and Vdh (50.3 kDa) were identified as responsible for the conversion of ferulic acid to vanillic acid; the proteins are encoded by the fca and vdh genes which are organized in an operon structure in the chromosome. The Vdh protein is 69% identical at the amino acid level to the Vdh protein recently identified in Pseudomonas sp. strain HR199 and converts vanillin to vanillic acid. Homology studies revealed that the Vdh proteins exhibited significant identity to aldehyde dehydrogenases from different organisms whereas Fca belonged to the enoyl-CoA hydratase family of proteins. Two proteins, designated VanA (39.9 kDa) and VanB (34.3 kDa), encoded by two genes, vanA and vanB, are organized in an operon in the chromosome. They were found to be responsible for the demethylation of vanillic acid to protocatechuic acid. The VanA proteins showed no homology to any other known protein, while VanB belonged to the ferredoxin family of proteins. This two-component enzyme system demethylated another phenolic monomer, veratric acid, thus indicating broad specificity. Studies of the regulation of the vanAB operon demonstrated that the genes were induced by the substrate, vanillic acid; however, the strongest induction was observed when cells were grown in the presence of the product of the reaction, protocatechuic acid.

Francesca Zennaro

Papers

Genetics of ferulic acid bioconversion to protocatechuic acid in plant-growth-promoting Pseudomonas putida WCS358