In situ proton NMR analysis of alpha-alkynoate biotransformations. From 'invisible' substrates to detectable metabolites.

Only 2% of the known natural products with acetylenic bonds are alpha-alkynoates. Their polarized, conjugated triple bond is an optimal target for an enzymic hydration. Therefore they are good substrates for the enzymes involved in metabolism of acetylenic compounds, resulting in products that are suitable for bacterial growth.

Acetylacetone-cleaving enzyme Dke1: a novel C-C-bond-cleaving enzyme from Acinetobacter johnsonii.

The toxicity of acetylacetone has been demonstrated in various studies. Little is known, however, about metabolic pathways for its detoxification or mineralization. Data presented here describe for the first time the microbial degradation of acetylacetone and the characterization of a novel enzyme that initiates the metabolic pathway.

A novel beta-diketone-cleaving enzyme from Acinetobacter johnsonii: acetylacetone 2,3-oxygenase.

A novel Fe+Zn containing oxygenase from Acinetobacter johnsonii catalyses 2,3-cleavage of acetylacetone to acetate and methylglyoxal has been purified. The stoichiometry of reactants and products conforms to a classical dioxygenase. The pure protein is a homotetramer of 64kD with variable amounts of Fe(2+) and Zn(2+).

In situ proton-NMR analyses of Escherichia coli HB101 fermentations in 1H2O and in D2O.

Experiments using one-dimensional Fourier-transform proton-NMR spectrometry for non-invasive analyses of microbial fermentations in situ, in vivo and in normal aqueous buffer are described. Analyses of the 'mixed acid' fermentation during growth of Escherichia coli on glucose and citrate were performed to identify and quantitatively estimate the concentrations of the two substrates provided and of the six products formed without sampling from the NMR tube. Identification of fermentation substrates and products was achieved by coincidence of selected diagnostic proton signals of individual compounds in the same solvent.