Tyrosine/dopa decarboxylase

FACCHINI, D., DELUCA, V., Phloem-specific expression of tyrosine dopa decarboxylase genes and the biosynthesis of isoquinoline alkaloids in opium poppy, Plant Cell, 1995, 7, 1811-1821. 

Isoquinoline Tyrosine/DOPA decarboxylase Papaver somniferum... 

FACCHINI, P.J., DE LUCA, V., Differential and tissue-specific expression of a gene family for tyrosine/dopa decarboxylase in opium poppy. J. Biol. Chem., 1994, 269,26684-26690. 

The benzylisoquinolines are formed from two molecules of fhe aromafic amino acid, tyrosine. In the past ten years, this pathway has been probed at the enzyme and gene level. The recent linking of the phloem-specific expression of tyrosine/Dopa decarboxylase (TYDC) genes with the bios)mthesis of the isoquinoline alkaloids in the opium poppy, Papaver somniferum (Facchini and De Luca, 1994, 1995, 2008 Liscombe and Facchini, 2008), and the association with alkaloid accumulation as part of the plant defence mechanism (Wink, 1993 Facchini et al, 1996) are of particular interest in furthering our knowledge of the location of alkaloid biosynthesis. 

IX. 4 -0-Methylation of 3-hydroxy-V-methylcoclaurine The several upstream biosynthetic genes in these steps, tyrosine/DOPA decarboxylase and tyrosine aminotransferase, have been isolated and characterized [13, 14]. 

Dopamine is a key intermediate in the plant BIA biosynthesis pathway. It condenses with 4-HPAA, and forms a BIA scaffold. In plants, tyrosine/ DOPA decarboxylases catalyze the decarboxylation of L-tyrosine and l-DOPA to tyramine and dopamine, respectively [49]. T3Tamine is an undesirable product for bacterial BIA synthetic pathways because its MAO product (i.e., 4-HPAA) combines with dopamine to form norcoclaurine, which needs CYP80B to be converted to reticuline. l-DOPA decarboxylase (PpDODC) from the Pseudomonas putida strain KT2440 exhibited a more than 10 -fold preference for l-DOPA compared with other aromatic amino acids [unpublished data]. Therefore, conversion of l-DOPA with PpDODC is expected to reduce the formation of undesirable by-products, 4-HPAA, and the resultant norcoclaurine (Fig. 1.4). Using an L-DOPA-produdng E. coli strain that overexpresses PpDODC, dopamine production reached 1.05 g. The conversion efficiency from L-tyrosine to dopamine was 29.1 % [24]. 

Figure 4.2 Biosynthesis of benzylisoquinoline alkaloids. Enzyme abbreviations TYDC, tyrosine/dopa decarboxylase NCS, norcoclaurine synth lse 60MT, norcoclaurine 6-O-methyltransferase 4 OMT, 3 -hydroxy-/V-methylcoclaurine 4 -0-methyltransferase CYP80A, berbamunine synth lse CYP80B, N-methylcoclaurine 3 -hydroxylase BBE, berberine bridge enzyme SOMT, scoulerine 9-0-methyltransferase CYP719A, (S)-canadine synthase SAT, salutaridinol 7-O-acetyltransferase COR, codeinone reductase.

The neurotransmitter must be present in presynaptic nerve terminals and the precursors and enzymes necessary for its synthesis must be present in the neuron. For example, ACh is stored in vesicles specifically in cholinergic nerve terminals. It is synthesized from choline and acetyl-coenzyme A (acetyl-CoA) by the enzyme, choline acetyltransferase. Choline is taken up by a high affinity transporter specific to cholinergic nerve terminals. Choline uptake appears to be the rate-limiting step in ACh synthesis, and is regulated to keep pace with demands for the neurotransmitter. Dopamine [51 -61-6] (2) is synthesized from tyrosine by tyrosine hydroxylase, which converts tyrosine to L-dopa (3,4-dihydroxy-L-phenylalanine) (3), and dopa decarboxylase, which converts L-dopa to dopamine. 

Fig. 2. Biosynthetic pathway for epinephrine, norepinephrine, and dopamine. The enzymes cataly2ing the reaction are (1) tyrosine hydroxylase (TH), tetrahydrobiopterin and O2 are also involved (2) dopa decarboxylase (DDC) with pyridoxal phosphate (3) dopamine-P-oxidase (DBH) with ascorbate, O2 in the adrenal medulla, brain, and peripheral nerves and (4) phenethanolamine A/-methyltransferase (PNMT) with. Cadenosylmethionine in the adrenal...

Together with dopamine, adrenaline and noradrenaline belong to the endogenous catecholamines that are synthesized from the precursor amino acid tyrosine (Fig. 1). In the first biosynthetic step, tyrosine hydroxylase generates l-DOPA which is further converted to dopamine by the aromatic L-amino acid decarboxylase ( Dopa decarboxylase). Dopamine is transported from the cytosol into synaptic vesicles by a vesicular monoamine transporter. In sympathetic nerves, vesicular dopamine (3-hydroxylase generates the neurotransmitter noradrenaline. In chromaffin cells of the adrenal medulla, approximately 80% of the noradrenaline is further converted into adrenaline by the enzyme phenylethanolamine-A-methyltransferase. 

Neural cells convert tyrosine to epinephrine and norepinephrine (Figure 31—5). While dopa is also an intermediate in the formation of melanin, different enzymes hydroxylate tyrosine in melanocytes. Dopa decarboxylase, a pyridoxai phosphate-dependent enzyme, forms dopamine. Subsequent hydroxylation by dopamine P-oxidase then forms norepinephrine. In the adrenal medulla, phenylethanolamine-A -methyltransferase uti-hzes S-adenosyhnethionine to methylate the primary amine of norepinephrine, forming epinephrine (Figure 31-5). Tyrosine is also a precursor of triiodothyronine and thyroxine (Chapter 42). 

By contrast, the cytoplasmic decarboxylation of dopa to dopamine by the enzyme dopa decarboxylase is about 100 times more rapid (Am 4x 10 " M) than its synthesis and indeed it is difficult to detect endogenous dopa in the CNS. This enzyme, which requires pyridoxal phosphate (vitamin B6) as co-factor, can decarboxylate other amino acids (e.g. tryptophan and tyrosine) and in view of its low substrate specificity is known as a general L-aromatic amino-acid decarboxylase. 

A glance at Fig. 15.4 will show that levodopa is metabolised primarily by dopa decarboxylase to DA and by COMT to 3-methoxy tyrosine, but usually referred to as OMD (o-methyldopa). 

Figure 2.16. Pathways for the synthesis and metabolism of the catecholamines. A=phenylalanine hydroxylase+pteridine cofactor+Oj B tyrosine hydroxylase+ tetrahydropteridme+Fe+ +Oj C=dopa decarboxylase+pyridoxal phosphate D= dopamine beta-oxidase+ascorbate phosphate+Cu+ +Oj E=phenylethanolamine N-methyltransferase+S-adenosylmethionine l=monoamine oxidase and aldehyde dehydrogenase 2=catechol-0-methyltransferase+S-adenosylmethionine.

Although numerous compounds have been tested as inhibitors of tyrosine hydroxylase 1379], no guanidines appear yet among them. Guanethidine itself has no significant effect on DOPA decarboxylase [323, 380] nor on dopamine-/3-oxidase [259,381,382]. Neither guanoxan, which is also a potent noradrenaline... 

Dopamine is the decarboxylation product of DOPA, dihydroxyphenylalanine, and is formed in a reaction catalysed by DOPA decarboxylase. This enzyme is sometimes referred to as aromatic amino acid decarboxylase, since it is relatively non-specific in its action and can catalyse decarboxylation of other aromatic amino acids, e.g. tryptophan and histidine. DOPA is itself derived by aromatic hydroxylation of tyrosine, using tetrahydrobiopterin (a pteridine derivative see Section 11.9.2) as cofactor. 

Synthesis of norepinephrine begins with the amino acid tyrosine, which enters the neuron by active transport, perhaps facilitated by a permease. In the neuronal cytosol, tyrosine is converted by the enzyme tyrosine hydroxylase to dihydroxyphenylalanine (dopa), which is converted to dopamine by the enzyme aromatic L-amino acid decarboxylase, sometimes termed dopa-decarboxylase. The dopamine is actively transported into storage vesicles, where it is converted to norepinephrine (the transmitter) by dopamine (3-hydroxylase, an enzyme within the storage vesicle. 

Noradrenergic neurons. The noradrenergic neuron uses NE for its neurotransmitter. Monoamine neurotransmitters are synthesized by means of enzymes, which assemble neurotransmitters in the cell body or nerve terminal. For the noradrenergic neuron, this process starts with tyrosine, the amino acid precursor of NE, which is transported into the nervous system from the blood by means of an active transport pump (Fig. 5 — 17). Once inside the neuron, the tyrosine is acted on by three enzymes in sequence, the first of which is tyrosine hydroxylase (TOH), the rate-limiting and most important enzyme in the regulation of NE synthesis. Tyrosine hydroxylase converts the amino acid tyrosine into dihydroxyphenylalanine (DOPA). The second enzyme DOPA decarboxylase (DDC), then acts, converting DOPA into dopamine (DA), which itself is a neurotransmitter in some neurons. However, for NE neurons, DA is just a precursor of NE. In fact, the third and final NE synthetic enzyme, dopamine beta-hydroxylase (DBH), converts DA into NE. The NE is then stored in synaptic packages called vesicles until released by a nerve impulse (Fig. 5—17). 

FIGURE 5—31. Dopamine (DA) is produced in dopaminergic neurons from the precursor tyrosine (tyr), which is transported into the neuron by an active transport pump, called the tyrosine transporter, and then converted into DA by two of the same three enzymes that also synthesize norepinephrine (Fig. 5-17). The DA-synthesizing enzymes are tyrosine hydroxylase (TOH), which produces DOPA, and DOPA decarboxylase (DDC), which produces DA. 

The general scheme of the biosynthesis of catecholamines was first postulated in 1939 (29) and finally confirmed in 1964 (Fig. 2) (30). Although not shown in Figure 2, in some cases the amino acid phenylalanine [63-91-2] can serve as a precursor it is converted in the liver to (-)-tyrosine [60-18-4] by the enzyme phenylalanine hydroxylase. Four enzymes are involved in E formation in the adrenal medulla and certain neurons in the brain tyrosine hydroxylase, dopa decarboxylase (also referred to as L-aromatic amino acid decarboxylase), dopamine-P-hydroxylase, and phenylethanolamine iV-methyltransferase. Neurons that form DA as their transmitter lack the last two of these enzymes, and sympathetic neurons and other neurons in the central nervous system that form NE as a transmitter do not contain phenylethanolamine N-methyl-transferase. The component enzymes and their properties involved in the formation of catecholamines have been purified to homogeneity and their properties examined. The human genes for tyrosine hydroxylase, dopamine- 3-oxidase and dopa decarboxylase, have been cloned (31,32). It is anticipated that further studies on the molecular structure and expression of these enzymes should yield interesting information about their regulation and function. 

Fig. 3. Schematic representation of the neurochemical events associated with neurotransmitter synthesis, release, re-uptake and metabolism in axons of diencephalic DA neurons terminating in classical synapses (Top Panel), and TIDA neurosecretory neurons terminating in close proximity to the hypophysial portal system (Botton Panel). Arrows with dashed lines represent end-product inhibition of TH activiy by DA (Top + Bottom Panels) or DA presynaptic autoreceptor-mediated inhibition of DA synthesis and release (Top Panel). Abbreviations COMT, Catechol-O-methyltransferase D, dopamine DDC, DOPA decarboxylase DOPA, 3,4-dihydrophenylalanine DOPAC, 3,4-dihydroxyphenylacetic acid HVA, homovanillic acid MAO, monoamine oxidase 3MT, 3-methoxytyramine TH, tyrosine hydroxylase.

Histamine, serotonin and the catecholamines (dopamine, epinephrine and norepinephrine) are synthesized from the aromatic amino acids histidine, tryptophan and phenylalanine, respectively. The biosynthesis of catecholamines in adrenal medulla cells and catecholamine-secreting neurons can be simply summarized as follows [the enzyme catalysing the reaction and the key additional reagents are in square brackets] phenylalanine — tyrosine [via liver phenylalanine hydroxylase + tetrahydrobiopterin] —> i.-dopa (l.-dihydroxyphenylalanine) [via tyrosine hydroxylase + tetrahydrobiopterin] —> dopamine (dihydroxyphenylethylamine) [via dopa decarboxylase + pyridoxal phosphate] — norepinephrine (2-hydroxydopamine) [via dopamine [J-hydroxylasc + ascorbate] —> epinephrine (jV-methyl norepinephrine) [via phenylethanolamine jV-methyltransferase + S-adenosylmethionine]. 

The standard assay contained retinal homogenate (0.2-3 mg protein), 0.05 mAf tyrosine, 1 mAf (6i ,S)-5,6,7,8-tetrahydro-L-biopterin, 3.5 mAf NADH, 0.02 unit of dihydroxypteridine reductase, 15 ng catalase, 40 mAf sodium acetate buffer (pH 6.0), and 0.1 mAf o-benzylhydroxylamine (inhibitor of Dopa decarboxylase) in a final volume of 100 fiL. After incubation of the mixture at 37°C for 5 to 20 minutes, the reaction as stopped by adding 100 fiL of 0.5 Af perchloric acid containing 0.4 mAf sodium metabisulfite and 0.1 mAf disodium EDTA. The supernate obtained after centrifugation was used for HPLC analysis. 

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