Publications

• Initial steps in the anaerobic degradation of 3,4,5-trihydroxybenzoate by Eubacterium oxidoreducens: characterization of mutants and role of 1,2,3,5-tetrahydroxybenzene.

  Haddock J.D., Ferry J.G.

  Chemical mutagenesis and antibiotic enrichment techniques were used to isolate five mutant strains of the obligate anaerobe Eubacterium oxidoreducens that were unable to grow on 3,4,5-trihydroxybenzoate (gallate). Two strains could not transform gallate and showed no detectable gallate decarboxyla ... >> More

  Chemical mutagenesis and antibiotic enrichment techniques were used to isolate five mutant strains of the obligate anaerobe Eubacterium oxidoreducens that were unable to grow on 3,4,5-trihydroxybenzoate (gallate). Two strains could not transform gallate and showed no detectable gallate decarboxylase activity. Two other strains transformed gallate to pyrogallol and dihydrophloroglucinol but lacked the hydrolase activity responsible for ring cleavage. A fifth strain accumulated pyrogallol, although it contained adequate levels of the enzymes proposed for the complete transformation of gallate to the ring cleavage product. The conversion of pyrogallol to phloroglucinol by cell extract of the wild-type strain was dependent on the addition of 1,2,3,5-tetrahydroxybenzene or dimethyl sulfoxide. This activity was induced by growth on gallate, while the other enzymes involved in the initial reactions of gallate catabolism were constitutively expressed during growth on crotonate. The results confirm the initial steps in the pathway previously proposed for the metabolism of gallate by E. oxidoreducens, except for the conversion of pyrogallol to phloroglucinol. << Less

  J Bacteriol 175:669-673(1993) [PubMed] [EuropePMC]

• Purification and properties of phloroglucinol reductase from Eubacterium oxidoreducens G-41.

  Haddock J.D., Ferry J.G.

  Phloroglucinol reductase was purified 90-fold to homogeneity from the anaerobic rumen organism Eubacterium oxidoreducens strain G-41. The enzyme is stable in the presence of air and is found in the soluble fraction after ultracentrifugation of cell extract. Ion-exchange, hydrophobic interaction, a ... >> More

  Phloroglucinol reductase was purified 90-fold to homogeneity from the anaerobic rumen organism Eubacterium oxidoreducens strain G-41. The enzyme is stable in the presence of air and is found in the soluble fraction after ultracentrifugation of cell extract. Ion-exchange, hydrophobic interaction, and affinity chromatography were used to purify the enzyme. The native Mr is 78,000, and the subunit Mr is 33,000 indicating an alpha 2 homodimer. The enzyme is specific for phloroglucinol and NADPH. The Km and Vmax are 600 microM and 640 mumol min-1 mg-1 (pH 7.2) for phloroglucinol, and 6.7 microM and 550 mumol min-1 mg-1 (pH 6.8) for NADPH; the Km and Vmax for the reverse direction are 290 microM and 140 mumol min-1 mg-1 (pH 7.2) for dihydrophloroglucinol, and 27 microM and 220 mumol min-1 mg-1 (pH 7.2) for NADP. Temperature and pH optima are 40 degrees C and 7.8 in the forward direction. The pure enzyme is colorless in solution and flavins are absent. Analysis for cobalt, manganese, molybdenum, vanadium, tungsten, selenium, copper, nickel, iron, and zinc indicated that these metals are not components of the phloroglucinol reductase. Cupric chloride, n-ethylmaleimide, and p-chloromercuribenzoate are potent inhibitors of enzyme activity. The properties of phloroglucinol reductase indicate that it functions in the pathway of anaerobic degradation of trihydroxybenzenes by catalyzing reduction of the aromatic nucleus prior to ring fission. << Less

  J. Biol. Chem. 264:4423-4427(1989) [PubMed] [EuropePMC]

• An aldolase-dependent phloroglucinol degradation pathway in <i>Collinsella</i> sp. zg1085.

  Li Y., Xu T., Tu Y., Li T., Wei Y., Zhou Y.

  Phloroglucinol (1,3,5-trihydroxybenzene) is a key intermediate in the degradation of polyphenols such as flavonoids and hydrolysable tannins and can be used by certain bacteria as a carbon and energy source for growth. The identification of enzymes that participate in the fermentation of phloroglu ... >> More

  Phloroglucinol (1,3,5-trihydroxybenzene) is a key intermediate in the degradation of polyphenols such as flavonoids and hydrolysable tannins and can be used by certain bacteria as a carbon and energy source for growth. The identification of enzymes that participate in the fermentation of phloroglucinol to acetate and butyrate in <i>Clostridia</i> was recently reported. In this study, we present the discovery and characterization of a novel metabolic pathway for phloroglucinol degradation in the bacterium <i>Collinsella</i> sp. zg1085, from marmot respiratory tract. In both the <i>Clostridial</i> and <i>Collinsella</i> pathways, phloroglucinol is first reduced to dihydrophoroglucinol by the NADPH-dependent phloroglucinol reductase (PGR), followed by ring opening to form (<i>S</i>)-3-hydroxy-5-oxohexanoate by a Mn²⁺-dependent dihydrophloroglucinol cyclohydrolase (DPGC). In the <i>Collinsella</i> pathway, (<i>S</i>)-3-hydroxy-5-oxohexanoate is then cleaved to form malonate semialdehyde and acetone by a newly identified aldolase (HOHA). Finally, a NADP⁺-dependent malonate-semialdehyde dehydrogenase converts malonate semialdehyde to CO₂ and acetyl-CoA, an intermediate in carbon and energy metabolism. Recombinant expression of the <i>Collinsella</i> PGR, DPGC, and HOHA in <i>E. coli</i> enabled the conversion of phloroglucinol into acetone, providing support for the proposed pathway. Experiments with <i>Olsenella profusa</i>, another bacterium containing the gene cluster of interest, show that the PGR, DPGC, HOHA, and MSDH are induced by phloroglucinol. Our findings add to the variety of metabolic pathways for the degradation of phloroglucinol, a widely distributed phenolic compound, in the anaerobic microbiome.IMPORTANCEPhloroglucinol is an important intermediate in the bacterial degradation of polyphenols, a highly abundant class of plant natural products. Recent research has identified key enzymes of the phloroglucinol degradation pathway in butyrate-producing anaerobic bacteria, which involves cleavage of a linear triketide intermediate by a beta ketoacid cleavage enzyme, requiring acetyl-CoA as a co-substrate. This paper reports a variant of the pathway in the lactic acid bacterium <i>Collinsella</i> sp. zg1085, which involves cleavage of the triketide intermediate by a homolog of deoxyribose-5-phosphate aldolase, highlighting the variety of mechanisms for phloroglucinol degradation by different anaerobic bacterial taxa. << Less

  Appl Environ Microbiol 90:e0104724-e0104724(2024) [PubMed] [EuropePMC]

  This publication is cited by 3 other entries.