Phenylethanolamine N-methyltransferase

Figure 42-10. Biosynthesis of catecholamines. (PNMT, phenylethanolamine-N-methyltransferase.)...

Because LCEC had its initial impact in neurochemical analysis, it is not, surprising that many of the early enzyme-linked electrochemical methods are of neurologically important enzymes. Many of the enzymes involved in catecholamine metabolism have been determined by electrochemical means. Phenylalanine hydroxylase activity has been determined by el trochemicaUy monitoring the conversion of tetrahydro-biopterin to dihydrobiopterin Another monooxygenase, tyrosine hydroxylase, has been determined by detecting the DOPA produced by the enzymatic reaction Formation of DOPA has also been monitored electrochemically to determine the activity of L-aromatic amino acid decarboxylase Other enzymes involved in catecholamine metabolism which have been determined electrochemically include dopamine-p-hydroxylase phenylethanolamine-N-methyltransferase and catechol-O-methyltransferase . Electrochemical detection of DOPA has also been used to determine the activity of y-glutamyltranspeptidase The cytochrome P-450 enzyme system has been studied by observing the conversion of benzene to phenol and subsequently to hydroquinone and catechol... 

Figure 1. Biosynthetic pathways for biogenic amines. In Drosophila and vertebrates decarboxylation of DOPA and 5-hydroxy-tryptophan is catalyzed by the same enzyme, DDC. In vertebrates this enzyme is called amino acid decarboxylase (AADC). Only vertebrates further metabolize dopamine to norepinephrine and epinephrine. TH, tryosine hydroxylase DDC, DOPA decarboxylase DBH, dopamine b-hydroxylase PNMT, phenylethanolamine N-methyltransferase. Tryp-OH tryptophan hydroxylase.

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.

In noradrenergic neurons, the end product is norepinephrine. In the adrenal medulla, the synthesis is carried one step further by the enzyme phenylethanolamine N-methyltransferase, which converts norepinephrine to epinephrine. The human adrenal medulla contains approximately four times as much epinephrine as norepinephrine. The absence of this enzyme in noradrenergic neurons accounts for the absence of significant amounts of epinephrine in noradrenergic neurons. The structures of these compounds are shown in Figure 9.4. 

Epinephrine is synthesized from NE in the adrenal medulla. Norepinephrine is methylated by phenylethanolamine-N-methyltransferase. Neurons containing this enzyme are also found in the CNS. 

Lipozits Zs, Phelix C, Pauli WK (1986) Electron microscopic analysis of tyrosine hydroxylase, dopamine-p-hydroxylase and phenylethanolamine-N-methyltransferase immunoreactive innervation of the hypothalamic paraventricular nucleus. Histochemistry 84 105 120. 

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 ... 

Norepinephrine diifuses into the cytosol, where it is converted to epinephrine by methylation of its amino group. This reaction, in which the methyl group is donated by S-adenosylmethionine, is catalyzed by S-adenosyl-L-methionine phenylethanolamine-N-methyltransferase (PNMT). Epinephrine enters the granules and remains there until it is released. 

Steps in the formation of classical neurotransmitters. AADC, amino acid decarboxylase AChE, acetylcholinesterase CAT, choline acetyltransferase COMT, catechol-O-methyltransfeiase DBH, dopamine P-hydroxylase DA, dopamine DOPA, dibydroxyphenylalanine GABA-T, GABA transaminase GAD, glutamic acid decarboxylase HD, histidine decarboxylase 5-HTP, 5-hydroxytrytophan MAO, monoamine oxidase PNMT, phenylethanolamine N-methyltransferase TH, tyrosine hydroxylase TPH, tryptophan hydroxylase. 

Figure 1. Pathway for the production of phenolamine from tyrosine. The reactions are catalyzed by the following enzymes DD, DOPA decarboxylase DBH, dopamine 8-hydroxylase PNMT, phenylethanolamine-N-methyltransferase. Data from reference 12.

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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