Human liver nicotinamide N-methyltransferase: ion-pairing radiochemical assay, biochemical properties and individual variation.
Nicotinamide N-methyltransferase (NNMT) catalyzes the N-methylation of nicotinamide, pyridine, and structurally related compounds. The products of this reaction are positively charged pyridinium ions that cannot be removed from aqueous solution by simple organic solvent extraction. We developed an assay for human liver NNMT based on the extraction of the positively charged reaction product into 60% isoamyl alcohol in toluene in the presence of the ion-pairing reagent 1-heptanesulfonic acid. Nicotinamide was the methyl acceptor for the reaction, and [14C-methyl]-S-adenosyl-L-methionine (Ado-Met) served as the methyl donor. Apparent Km values of human liver NNMT for nicotinamide and Ado-Met were 347 and 1.76 mumol/l, respectively. The product of the reaction was identified as N1-methylnicotinamide (NMN) by high performance liquid chromatography. NNMT activity was inhibited by the reaction products, NMN and S-adenosyl-L-homocysteine. NNMT activity was not affected by inhibitors of other methyltransferases including Ca2+, SKF 525A and 3,4-dimethoxy-5-hydroxybenzoic acid. Individual variation in NNMT activity was studied by measuring hepatic enzyme activities in 163 human liver biopsy samples obtained during clinically-indicated surgery. The average NNMT activity in these samples was 51.5 +/- 32.5 U per mg protein (mean +/- SD), and there was not a significant correlation of enzyme activity with patient age or with the time of storage of the biopsy samples at -80 degrees C. The distribution of activities was bimodal, and approximately 26% of the samples were included in a subgroup with high NNMT activity. It will now be possible to test the hypothesis that individual differences in hepatic NNMT activity might be related to variation in the N-methylation of pyridine compounds and to individual differences in either toxicity or the therapeutic efficacy of such compounds.
Rini
J
J
Department of Pharmacology, Mayo Clinic, Rochester, MN 55905.
Szumlanski
C
C
Guerciolini
R
R
Weinshilboum
R M
RM
D006801:Humans; D008099:Liver / Q000201:enzymology*; D008780:Methyltransferases / Q000378:metabolism*; D050903:Nicotinamide N-Methyltransferase; D011854:Radiochemistry
D008780:Methyltransferases; C494406:NNMT protein, human; D050903:Nicotinamide N-Methyltransferase