This enzyme is one of the three enzymes involved in L-dopa (3,4-dihydroxy-L-phenylalanine) catabolism in the non-oxygenic phototrophic bacterium Rubrivivax benzoatilyticus OU5 (and not Rhodobacter sphaeroides OU5 as had been thought ), the other two being EC 4.3.1.22 (dihydroxyphenylalanine reductive deaminase) and EC 2.6.1.49 (3,4-dihydroxyphenylalanine transaminase). In addition to L-dopa, the enzyme can also use L-tyrosine, L-phenylalanine, L-tryptophan and glutamate as substrate, but more slowly. The enzyme is inhibited by NADH and 2-oxoglutarate.
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota
This enzyme is one of the three enzymes involved in L-dopa (3,4-dihydroxy-L-phenylalanine) catabolism in the non-oxygenic phototrophic bacterium Rubrivivax benzoatilyticus OU5 (and not Rhodobacter sphaeroides OU5 as had been thought [1]), the other two being EC 4.3.1.22 (dihydroxyphenylalanine reductive deaminase) and EC 2.6.1.49 (3,4-dihydroxyphenylalanine transaminase). In addition to L-dopa, the enzyme can also use L-tyrosine, L-phenylalanine, L-tryptophan and glutamate as substrate, but more slowly. The enzyme is inhibited by NADH and 2-oxoglutarate.
Substrates: oxidative deamination, unusual oxygen-consuming reaction catalyzed by the enzyme toward aromatic amines (serotonin, dopamine, and alpha-methyldopamine) and D-tryptophan methyl ester Products: production in equivalent amounts depending on the nature of the substrate, and ammonia with concomitant O2 consumption in a 1:2 molar ratio with respect to the products. A ketimine accumulates during the linear phase of product formation. This species is reactive since it is converted back to pyridoxal 5'-phosphate when the substrate is consumed. Superoxide anion and hydrogen peroxide are both generated during the catalytic cycles.
Substrates: production depending on the nature of the substrate, and ammonia with concomitant O2 consumption in a 1:2 molar ratio with respect to the products Products: -
Substrates: The novelty in DDC is the possibility of catalyzing a reaction involving dioxygen although the enzyme lacks of any cofactor or metal related to O2 chemistry. The external aldimine intermediate undergoes a decarboxylation or a deprotonation leading to a quinonoid species, that is protonated at C4 producing the ketimine intermediate. Although it cannot be ruled out that this intermediate could be attacked by dioxygen, it seems much more likely, regarding enzymes proceeding through a carbanion chemistry on DDC, that the more electron dense quinonoid intermediate, in equilibrium with the ketimine, is reactive toward O2. Aerobiosis shifts the quinonoid-ketimine equilibrium toward quinonoid, while anaerobiosis shifts the equilibrium toward ketimine. The reaction between dioxygen and the quinonoid give rise directly to a superoxide anion and semiquinone. Superoxide is deprotonated and its anionic form is thus able to couple with the semiquinone giving rise to a peroxide species that is further protonated, and thus forming a hydroperoxy-pyridoxal 5'-phosphate intermediate. This rearranges to produce aldehyde, ammonia and hydrogen peroxide. Products: -
Substrates: oxidative deamination, unusual oxygen-consuming reaction catalyzed by the enzyme toward aromatic amines (serotonin, dopamine, and alpha-methyldopamine) and D-tryptophan methyl ester Products: production in equivalent amounts depending on the nature of the substrate, and ammonia with concomitant O2 consumption in a 1:2 molar ratio with respect to the products. A ketimine accumulates during the linear phase of product formation. This species is reactive since it is converted back to pyridoxal 5'-phosphate when the substrate is consumed. Superoxide anion and hydrogen peroxide are both generated during the catalytic cycles.
Substrates: production depending on the nature of the substrate, and ammonia with concomitant O2 consumption in a 1:2 molar ratio with respect to the products Products: -
Substrates: The novelty in DDC is the possibility of catalyzing a reaction involving dioxygen although the enzyme lacks of any cofactor or metal related to O2 chemistry. The external aldimine intermediate undergoes a decarboxylation or a deprotonation leading to a quinonoid species, that is protonated at C4 producing the ketimine intermediate. Although it cannot be ruled out that this intermediate could be attacked by dioxygen, it seems much more likely, regarding enzymes proceeding through a carbanion chemistry on DDC, that the more electron dense quinonoid intermediate, in equilibrium with the ketimine, is reactive toward O2. Aerobiosis shifts the quinonoid-ketimine equilibrium toward quinonoid, while anaerobiosis shifts the equilibrium toward ketimine. The reaction between dioxygen and the quinonoid give rise directly to a superoxide anion and semiquinone. Superoxide is deprotonated and its anionic form is thus able to couple with the semiquinone giving rise to a peroxide species that is further protonated, and thus forming a hydroperoxy-pyridoxal 5'-phosphate intermediate. This rearranges to produce aldehyde, ammonia and hydrogen peroxide. Products: -
carbiDOPA, addition of 10 microM inhibitor to reaction mixtures (Y332F mutant with L-dopa) in the presence or in the absence of catalase or superoxide dismutase, immediately stops the O2 consumption.
Y332F DDC mutant, reaction in 50 mM Hepes, pH 7.5, at 25°C causes the production of ammonia and 3,4-dihydroxyphenylacetaldehyde along with the consumption of molecular oxygen in a 1:2 molar ratio
mutant T246A presents a decarboxylase activity, the kcat value is decreased by 29fold with respect to wild-type, an oxidative deamination reaction of aromatic amines does not occur
T246A mutant catalyzes the oxidative deamination of D-tryptophan methyl ester with a kcat value approximately 5% with respect to the measured value for wild-type
wild-type enzyme and Y332F variant are able to perform the oxidation toward aromatic amines or aromatic L-amino acids, without the aid of any cofactor related to oxygen chemistry.
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