L-2,3-diaminopropionate generates diverse metabolic stresses in Salmonella enterica.

Unchecked amino acid accumulation in living cells has the potential to cause stress by disrupting normal metabolic processes. Thus, many organisms have evolved degradation strategies that prevent endogenous accumulation of amino acids. L-2,3-diaminopropionate (Dap) is a non-protein amino acid produced in nature where it serves as a precursor to siderophores, neurotoxins and antibiotics. Dap accumulation in Salmonella enterica was previously shown to inhibit growth by unknown mechanisms. The production of diaminopropionate ammonia-lyase (DpaL) alleviated Dap toxicity in S. enterica by catalyzing the degradation of Dap to pyruvate and ammonia. Here, we demonstrate that Dap accumulation in S. enterica elicits a proline requirement for growth and specifically inhibits coenzyme A and isoleucine biosynthesis. Additionally, we establish that the DpaL-dependent degradation of Dap to pyruvate proceeds through an unbound 2-aminoacrylate (2AA) intermediate, thus contributing to 2AA stress inside the cell. The reactive intermediate deaminase, RidA, is shown to prevent 2AA damage caused by DpaL-dependent Dap degradation by enhancing the rate of 2AA hydrolysis. The results presented herein inform our understanding of the effects Dap has on metabolism in S. enterica, and likely other organisms, and highlight the critical role played by RidA in preventing 2AA stress stemming from Dap detoxification.

Electron spin-echo studies of the composition of the paramagnetic intermediate formed during the deamination of propanolamine by ethanolamine ammonia-lyase, and AdoCbl-dependent enzyme.

During the deamination of S-2-aminopropanol by the AdoCbl-dependent ethanolamine ammonia-lyase of Clostridia sp., a catalytic intermediate accumulates whose active site contains two paramagnetic species: cob(II)alamin and a free radical derived from the substrate molecule. Spin-echo spectroscopy has revealed that the unpaired electron on the substrate-derived radical is delocalized over a nitrogen atom that from its quadrupole splittings is probably a component of a secondary amide group. Experiments with 15N- and deuterium-labeled propanolamine gave no evidence of an interaction between this unpaired electron and the nitrogen originally attached to the substrate molecule. These results strongly suggest that the substrate-derived radical in this intermediate has already lost its nitrogen, and that this radical is stabilized by delocalization of the unpaired electron onto a nitrogen most likely situated in one of the peptide bonds of the enzyme backbone.

Phenylalanine ammonia lyase in the management of phenylketonuria: the relationship between ingested cinnamate and urinary hippurate in humans.

The hypothesis that the action of an oral preparation of the enzyme phenylalanine ammonia lyase, used to lower blood phenylalanine levels in phenylketonuria, is by removal of phenylalanine from hydrolysed protein in the lumen of the gut has been examined. Phenylalanine ammonia lyase mediates the conversion of phenylalanine to cinnamate. If cinnamate is produced in the gut it will be absorbed and converted to hippurate in the liver. However, treatment with phenylalanine ammonia lyase does not result in raised urinary hippurate ammonia lyase does not result in raised urinary hippurate levels while ingestion of cinnamate does.

Biochemical properties and immunogenicity of L-phenylalanine ammonia-lyase: effects on tumor-bearing mice.

L-Phenylalanine ammonia-lyase (PAL) from yeast was used to deplete plasma L-phenylalanine and L-tyrosine in an attempt to achieve inhibition of tumor growth in mice. Plasma L-phenylalanine and L-tyrosine were reduced to nondetectable levels when circulating PAL activity was maintained at greater than or equal to 0.06 unit/ml. Repeated administration resulted in the appearance of anti-PAL antibodies. A radioimmunoassay based on the method of Farr was developed to determine quantitatively the presence of anti-PAL. Sublethal total-body irradiation temporarily suppressed the immunologic response of the host. Long-term specific immunosuppression to PAL was achieved with cyclophosphamide (CPA). A single dose of 180 mg/kg of CPA administered ip to mice 24 hours before, simultaneously with, or 24 hours after 100 units/kg of PAL induced tolerance for 450 days (20 injections of enzyme). The plasma half-life of PAL in CPA-treated mice remained essentially the same as that found after a single injection (25 hours), and anti-PAL probably will require specific immunosuppression of the host to repeated injections of the enzyme.