Phenylethanolamine ,A-methyltransferase

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. 

Trace Amines. Figure 1 The main routes of trace amine metabolism. The trace amines (3-phenylethylamine (PEA), p-tyramine (TYR), octopamine (OCT) and tryptamine (TRP), highlighted by white shading, are each generated from their respective precursor amino acids by decarboxylation. They are rapidly metabolized by monoamine oxidase (MAO) to the pharmacologically inactive carboxylic acids. To a limited extent trace amines are also A/-methylated to the corresponding secondary amines which are believed to be pharmacologically active. Abbreviations AADC, aromatic amino acid decarboxylase DBH, dopamine b-hydroxylase NMT, nonspecific A/-methyltransferase PNMT, phenylethanolamine A/-methyltransferase TH, tyrosine hydroxylase. 

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). 

Tyrosine is converted to dopa by the rate-limiting enzyme tyrosine hydroxylase, which requires tetrahydrobiopterin, and is inhibited by a-methyltyrosine. Dopa is decarboxylated to dopamine by L-aromatic amino acid decarboxylase, which requires pyridoxal phosphate (vitamin B6) as a coenzyme. Carbidopa, which is used with levodopa in the treatment of parkinsonism, inhibits this enzyme. Dopamine is converted to norepinephrine by dopamine P-hydroxylase, which requires ascorbic acid (vitamin C), and is inhibited by diethyldithiocarbamate. Norepinephrine is converted to epinephrine by phenylethanolamine A -methyltransferase (PNMT), requiring S-adeno-sylmethionine. The activity of PNMT is stimulated by corticosteroids. 

Figure 7.1 Pathways of synthesis and metabolism of catecholamines with enzymes catalyzing various reactions. (1) Tyrosine hydroxylase (2) aromatic amino acid decarboxylase (3) phenylamine-P-hydroxylase (4) phenylethanolamine-A-methyltransferase (5) monoamine oxidase plus aldehyde dehydrogenase (6) catechol-O-methyltransferase (7) conjugating enzymes about 95% phenolsulfo-transferase and 5% phenolglucuronatetransferase (in human). DOPA, dihydroxyphenylalanine DOMA, dihydroxymandelic acid DHPG, dihydroxyphenylglycol DOPAC, dihydroxyphenylacetic acid HVA, homovanillic acid MHPG, methoxyhydroxylphenylglycol VMA, vanilmandelic acid...

The enzymes involved in the formation of the catecholamines are of low specificity. DOPA decarboxylase, or an enzyme closely akin to it, is concerned in the formation of 5-hydroxytryptamine > dopamine-/9-oxidase has been shown to be capable of hydroxylating the jd-carbon atom of a number of tyramine derivatives - and phenylethanolamine A-methyltransferase is equally unselective in its A-methylation of noradrenaline derivatives . This lack of specificity suggests the possibility that alternative pathways for the formation of noradrenaline and adrenaline might exist in vivo. Some of the putative intermediaries in these other pathways have been shown to occur naturally and one of them, octopamine (/) is found in the brain . 

B. Catecholamines Dopa decarboxylase Dopamine hydroxylase Phenylethanolamine A -methyltransferase Monoamine oxidase... 

Borchardt, R. T., Vincek, W. C., and Grunewald, G. L., 1977, A liquid chromatographic assay for phenylethanolamine-A-methyltransferase, Anal. Biochem. 82 149-157. 

In cells that synthesize epinephrine, the final step in the pathway is catalyzed by the enzyme phenylethanolamine /V-methyltransferase. This enzyme is found in a small group of neurons in the brainstem that use epinephrine as their neurotransmitter and in the adrenal medullary cells, for which epinephrine is the primary hormone secreted. Phenylethanolamine N-methyltransferase (PNMT) transfers a methyl group from S-adenosylmethionine to the nitrogen of norepinephrine, forming a secondary amine [5]. The coding sequence of bovine PNMT is contained in a... 

Baetge, E. E., Suh, Y. H. and Joh, T. H. Complete nucleotide and deduced amino acid sequence of bovine phenylethanolamine N-methyltransferase partial amino acid homology with rat tyrosine hydroxylase. Proc. Natl Acad. Sci. U.S.A. 83 5454-5458,1986. 

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.

This enzyme [EC 2.1.1.28], also known as phenylethanol-amine A -methyltransferase, catalyzes the reaction of S-adenosyl-L-methionine with phenylethanolamine to produce 5-adenosyl-L-homocysteine and A -methylphenyl-ethanolamine. The enzyme will act on a number of phe-nylethanolamines and will catalyze the conversion of noradrenalin (or norepinephrine) into adrenalin (or epinephrine). 

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. 

A series of tetrahydrothiadiazoloisoquinolines that could potentially reduce the formation of epinephrine by inhibiting phenylethanolamine-jV-methyltransferase has been prepared <89JMC1566>. 

In the assay described by Beaudouin et al. (1993), the phenylethanolamine AT-methyltransferase catalyzes the transfer of a methyl group from S-adenosyl-L-methionine to noradrenaline to form adrenaline and S-adenosyl-L-homocys-teine as the final step of adrenaline biosynthesis. Adrenaline is mainly synthesized in the adrenal medulla. 

Figure 9.14 Typical elution pattern of phenylethanolamine N-methyltransferase incubation mixtures with the homogenate of rat pons plus medulla oblongata as enzyme. The incubation mixture contained 10 mg of rat pons plus medulla oblongata as enzyme and 16 fxM noradrenaline (NA) and 18 fxM S-adenosyl-L-methionine (SAM) as substrates. (A) Experimental incubation with homogenate of 10 mg of rat pons plus medulla oblongata. (B) Blank incubation without enzyme. (C) Another blank incubation, to which was added IS pmol of adrenaline (AD) the reaction had been stopped. Formation of 16. 6 pmol of AD from NA during 60 minutes of incubation at 37°C was calculated from a calibration curve. DHBA, dihydroxybenzylamine (internal standard) UN, unknown peak. (From Trocewicz et al., 1982.)...

The additional presence of phenylethanolamine N-methyltransferase in adrenal medullary chromaffin cells leads to further conversion of norepinephrine to epinephrine (Figure 29-2). Since phenylethanolamine N-methyitransferase is a cytosolic enzyme, this step depends on leakage of norepinephrine from vesicular storage granules into the ceU cytoplasm and the transfer of a methyl group from S-adenosylmethionine to norepinephrine. Epinephrine is then translocated into chromaffin granules where the amine is stored, awaiting release. 

The free 0-methylated amine metabolites are present in plasma at picomolar concentrations that have made their accurate measurement technically difficult. Measurements of plasma metanephrines therefore represent relatively recent developments. The.first method enabling accurate measurement of plasma free normetanephrine involved a radioenzymatic assay in which normetanephrine was converted to H-iabeled metanephrine using preparations of the enzyme phenylethanolamine-N-methyltransferase, incubated with H-methyi-labeled S-adenosylmethionine. This method, however, did not allow measurements of metanephrine or methoxytyramine, and therefore had limited clinical utility. 

Levels of phenylethanolamine-N-methyltransferase (PNMT) have been found to be significantly raised in discrete brain stem regions in both SHR and DOCA-salt hypertensive rats.33 Administration of SKF 7698 (7), a PNMT inhibitor, normalized pressure in DOCA rats. The possibility that the effect was due to the adrenolytic action of (7) was not excluded. The best of a new series of PNMT inhibitors with much reduced a-blocking properties is SKF 6 139 (8).3 ... 

As described, the secretion of epinephrine in response to stress, trauma, extreme exercise, or hypoglycemia causes a rapid mobilization of energy stores, that is, glucose from the liver and fatty acids from adipose tissue. The reaction in which NE is methylated to form E is mediated by the enzyme phenylethanolamine-N-methyltransferase (PNMT). Although the enzyme occurs predominantly in the chromaffin cells of the adrenal medulla, it is also... 

Monoamine oxidase is a flavoprotein that catalyzes the oxidative deamination of amines to form the corresponding aldehydes. 02 is the electron acceptor, and NH3 and H202 are the other products. (PNMT = phenylethanolamine-N-methyltransferase.)... 

Transferases are enzymes that catalyze the transfer of functional groups from one molecule to another. For example, a transaminase catalyzes the transfer of an amino functional group, and a kinase catalyzes the transfer of a phosphate group. Kinases play a major role in energy-harvesting processes involving ATP. In the adrenal glands, norepinephrine is converted to epinephrine by the enzyme phenylethanolamine-N-methyltransferase (PNMT), a transmethylase. 

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