Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:4.1.2.13 (aldolase)
 3,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Pseudomonas putida KT2440 is a physiologically extremely versatile non-pathogenic bacterium that is applied as a "biosafety strain" in biotechnological processes, as authorized by the USA National Institute of Health. Analysis of the P. putida KT2440 whole-genome sequence revealed the genetic organization of the genes fcs, ech, and vdh, which are essential for ferulic acid conversion to vanillic acid via vanillin. To confirm the physiological function of these structural genes as feruloyl-CoA synthetase (Fcs), enoyl-CoA hydratase/aldolase (Ech), and vanillin dehydrogenase (Vdh), respectively, they were cloned and expressed in Escherichia coli. Recombinant strains harboring fcs and ech were able to transform ferulic acid to vanillin. The enzyme activities of Fcs and Vdh were determined in protein extracts of these cells. The essential involvement of fcs, ech and vdh in the catabolism of ferulic acid in P. putida KT2440 was proven by separately inactivating each gene by insertion of Omega-elements. The corresponding mutant strains KT2440 fcsOmegaKm, KT2440 echOmegaKm, and KT2440 vdhOmegaKm were not able to grow on ferulic acid. The potential application of P. putida KT2440 and the mutant strains in biotechnological vanillin production process is discussed.
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PMID:Functional analyses of genes involved in the metabolism of ferulic acid in Pseudomonas putida KT2440. 1276 69

The potential of two Rhodococcus strains for biotechnological vanillin production from ferulic acid and eugenol was investigated. Genome sequence data of Rhodococcus sp. I24 suggested a coenzyme A-dependent, non-beta-oxidative pathway for ferulic acid bioconversion, which involves feruloyl-CoA synthetase (Fcs), enoyl-CoA hydratase/aldolase (Ech), and vanillin dehydrogenase (Vdh). This pathway was proven for Rhodococcus opacus PD630 by physiological characterization of knockout mutants. However, expression and functional characterization of corresponding structural genes from I24 suggested that degradation of ferulic acid in this strain proceeds via a beta-oxidative pathway. The vanillin precursor eugenol facilitated growth of I24 but not of PD630. Coniferyl aldehyde was an intermediate of eugenol degradation by I24. Since the genome sequence of I24 is devoid of eugenol hydroxylase homologous genes (ehyAB), eugenol bioconversion is most probably initiated by a new step in this bacterium. To establish eugenol bioconversion in PD630, the vanillyl alcohol oxidase gene (vaoA) from Penicillium simplicissimum CBS 170.90 was expressed in PD630 together with coniferyl alcohol dehydrogenase (calA) and coniferyl aldehyde dehydrogenase (calB) genes from Pseudomonas sp. HR199. The recombinant strain converted eugenol to ferulic acid. The obtained data suggest that genetically engineered strains of I24 and PD630 are suitable candidates for vanillin production from eugenol.
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PMID:Potential of Rhodococcus strains for biotechnological vanillin production from ferulic acid and eugenol. 1642 16

Vanillin is one of the most important flavors in the food industry and there is great interest in its production through biotechnological processes starting from natural substrates such as ferulic acid. Among bacteria, recombinant Escherichia coli strains are the most efficient vanillin producers, whereas Pseudomonas spp. strains, although possessing a broader metabolic versatility, rapidly metabolize various phenolic compounds including vanillin. In order to develop a robust Pseudomonas strain that can produce vanillin in high yields and at high productivity, the vanillin dehydrogenase (vdh)-encoding gene of Pseudomonas fluorescens BF13 strain was inactivated via targeted mutagenesis. The results demonstrated that engineered derivatives of strain BF13 accumulate vanillin if inactivation of vdh is associated with concurrent expression of structural genes for feruloyl-CoA synthetase (fcs) and hydratase/aldolase (ech) from a low-copy plasmid. The conversion of ferulic acid to vanillin was enhanced by optimization of growth conditions, growth phase and parameters of the bioconversion process. The developed strain produced up to 8.41 mM vanillin, which is the highest final titer of vanillin produced by a Pseudomonas strain to date and opens new perspectives in the use of bacterial biocatalysts for biotechnological production of vanillin from agro-industrial wastes which contain ferulic acid.
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PMID:Metabolic engineering of Pseudomonas fluorescens for the production of vanillin from ferulic acid. 2187 27

Vanillin is one of the most important flavoring agents used today. That is why many efforts have been made on biotechnological production from natural abundant substrates. In this work, the nonpathogenic Pseudomonas putida strain KT2440 was genetically optimized to convert ferulic acid to vanillin. Deletion of the vanillin dehydrogenase gene (vdh) was not sufficient to prevent vanillin degradation. Additional inactivation of a molybdate transporter, identified by transposon mutagenesis, led to a strain incapable to grow on vanillin as sole carbon source. The bioconversion was optimized by enhanced chromosomal expression of the structural genes for feruloyl-CoA synthetase (fcs) and enoyl-CoA hydratase/aldolase (ech) by introduction of the strong tac promoter system. Further genetic engineering led to high initial conversion rates and molar vanillin yields up to 86% within just 3 h accompanied with very low by-product levels. To our knowledge, this represents the highest productivity and molar vanillin yield gained with a Pseudomonas strain so far. Together with its high tolerance for ferulic acid, the developed, plasmid-free P. putida strain represents a promising candidate for the biotechnological production of vanillin.
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PMID:Genetic engineering of Pseudomonas putida KT2440 for rapid and high-yield production of vanillin from ferulic acid. 2413 72

Using genome-wide mutant fitness assays in diverse bacteria, we identified novel oxidative pathways for the catabolism of 2-deoxy-d-ribose and 2-deoxy-d-ribonate. We propose that deoxyribose is oxidized to deoxyribonate, oxidized to ketodeoxyribonate, and cleaved to acetyl coenzyme A (acetyl-CoA) and glyceryl-CoA. We have genetic evidence for this pathway in three genera of bacteria, and we confirmed the oxidation of deoxyribose to ketodeoxyribonate in vitro. In Pseudomonas simiae, the expression of enzymes in the pathway is induced by deoxyribose or deoxyribonate, while in Paraburkholderia bryophila and in Burkholderia phytofirmans, the pathway proceeds in parallel with the known deoxyribose 5-phosphate aldolase pathway. We identified another oxidative pathway for the catabolism of deoxyribonate, with acyl-CoA intermediates, in Klebsiella michiganensis. Of these four bacteria, only P. simiae relies entirely on an oxidative pathway to consume deoxyribose. The deoxyribose dehydrogenase of P. simiae is either nonspecific or evolved recently, as this enzyme is very similar to a novel vanillin dehydrogenase from Pseudomonas putida that we identified. So, we propose that these oxidative pathways evolved primarily to consume deoxyribonate, which is a waste product of metabolism. IMPORTANCE Deoxyribose is one of the building blocks of DNA and is released when cells die and their DNA degrades. We identified a bacterium that can grow with deoxyribose as its sole source of carbon even though its genome does not contain any of the known genes for breaking down deoxyribose. By growing many mutants of this bacterium together on deoxyribose and using DNA sequencing to measure the change in the mutants' abundance, we identified multiple protein-coding genes that are required for growth on deoxyribose. Based on the similarity of these proteins to enzymes of known function, we propose a 6-step pathway in which deoxyribose is oxidized and then cleaved. Diverse bacteria use a portion of this pathway to break down a related compound, deoxyribonate, which is a waste product of metabolism. Our study illustrates the utility of large-scale bacterial genetics to identify previously unknown metabolic pathways.
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PMID:Oxidative Pathways of Deoxyribose and Deoxyribonate Catabolism. 3074 95