Figure 1: Conversion of norepinephrine to epinephrine by Phenylethanolamine N-Methyltransferase in the adreal medulla (from The Bello Lectures)
Phenylethanolamine N-Methyltransferase
Gene Map and Alignment
Figure 2: Gene map of human PNMT (from Kepp et al. (2007); reference 5)The human PNMT has been mapped to chromosome 17q22-q24 and consists of three exons and two introns spanning about 2100 basepairs [1]. This gene encodes a protein with 282 amino acid residues and a predicted molecular weight of 30 853 kDa, including the initial methionine. Interestingly, this amino acid sequence was 88% homologous to that of bovine PNMT [1, 2].
Figure 3: Alignment of Sus (Pig), Danio (Zebrafish), Homo (Human) and Mus (Mouse) PNMT (generated by ClustalW)Key (from ClustalW):
"*" means that the residues or nucleotides in that column are identical in all sequences in the alignment.
":" means that conserved substitutions have been observed.
"." means that semi-conserved substitutions are observed.
Color Key (from ClustalW):
| AVFPMILW | RED | Small (small+ hydrophobic (incl.aromatic -Y)) |
| DE | BLUE | Acidic |
| RK | MAGENTA | Basic |
| STYHCNGQ | GREEN | Hydroxyl + Amine + Basic - Q |
| Others | Gray |
Figure 4: Cladogram showing relationship between PNMT of pig, human, mouse, and zebrafish (generated by ClustalW)
Figure 5: Score table of alignment. Note that the higher the score, the higher the similarity between proteins. From this table, human and mouse PNMT are the most similar. (Generated from ClustalW)Active Site
PNMT converts norepinephrine to epinephrine by transferring a methyl group from S-adenosyl-L-methionine to the primary amine of norepinephrine to form epinephrine and the cofactor product S-adenosyl-L-homocysteine [2, 4, 5]. X-ray structures show that the active site of PNMT is covered and a significant conformational change is required to make the active site accessible for the binding of substrates [4, 5]. Several amino acid residues have been discovered to be responsible for substrate positioning and binding, including Asp267 which is important in positioning the substrate but does not participate directly in catalysis [3, 5]. This amino acid presumably works through its interaction with the side-chain hydroxyl of the phenylethanolamines. Experimental results have shown S-adenosyl-L-methionine causes the conformational change necessary for conversion of norepinephrine to epinephrine [2, 3, 5].
Figure 6: Human PNMT amino acids (in yellow) interacting with an inhibitor (the white central molecule) at the active site (From Grunewald et al. (2007); Reference 3)References
[1] Baetge, E. E., Behringer, R. R., Messing, A., Brinster, R. L., Palmiter, R. D. (1988). Transgenic mice express the human phenylethanolamine N-methyltransferase gene in adrenal medulla and retina. Proc. Nat. Acad. Sci. 85: 3648-3652
[2] Goldstein, D. S., Eisenhofer, G., and Kopin, I. J. (2006). Clinical Catecholamine Neurochemistry: A Legacy of Julius Axelrod. Cellular and Molecular Neurobiology. 26: 695-702
[3] Grunewald, G. L., Seim, M. R., Regier, R. C., and Criscione, K. R. (2007). Exploring the Active Site of Phenylethanolamine N-Methyltransferase with 1,2,3,4-Tetrahydrobenz[h]isoquinoline Inhibitors. Bioorg Med Chem. 15(3): 1298-1310
[4] Hilbert, P., Lindpaintner, K., Beckmann, J. S., Serikawa, T., Soubrier, F., Dubay, C., Cartwright, P., De Gouyon, B., Julier, C., Takahasi, S., Vincent, M., Ganten, D., Georges, M., Lathrop, G. M. (1991). Chromosomal mapping of two genetic loci associated with blood-pressure regulation in hereditary hypertensive rats. Nature. 353: 521-529
[5] Kepp, K., Juhanson, P., Kozich, V., Ots, M., Viigimaa, M., and Laan, M. (2007). Resequencing PNMT in European hypertensive and normotensive individuals: no common susceptibility variants for hypertension and purifying selection on intron I. BMC Medical Genetics. 8:(47)
[6] Norris, David O. (2007). Vertebrate Endocrinology. Burlington, MA, USA: Elsevier Academic Press.
[7] Silverthorn, D. (2007). Human Physiology: An Integrated Approach. San Francisco, USA: Benjamin Cummings.
[8] Stolk, Jon M., U’Prichard, David C., Fuxe, Kjell. (Eds.). (1988). Epinephrine in the Central Nervous System. Oxford: Oxford University Press.
