A radiochemical assay for the measurement of histamine N-methyltransferase (HNMT) activity in human erythrocytes (RBCs) has been developed. This assay was developed as a first step toward testing the hypothesis that the biochemical properties and regulation of HNMT in an easily obtainable human cell, the RBC, might reflect those of the enzyme in less accessible cells and tissues. The Michaelis (Km) constant in the RBC for histamine, the methyl acceptor substrate for the reaction, was 5.0 × 10-5 mol/l. The Km constant for S-adenosyl-l-methionine, the methyl donor, was 2.8 × 10-6 mol/l. The assay was performed at a reaction pH of 7.4 with a potassium phosphate buffer. The product of the reaction was identified as NT-methylhistamine by high performance liquid chromatography. The Kii for inhibition of the RBC enzyme by amodiaquine, an HNMT inhibitor, was 1.O × 10-7 mol/l, while the Kis value was 0.48 × 10-7 mol/l. Blood samples obtained from 39 randomly selected adult white subjects had a mean activity of 130 ± 30 U/ml of packed RBCs (mean ± SD). Enzyme activities varied over a range from 74-213 U. There were no differences between men and women in mean activities, nor was there a significant correlation between RBC HNMT activity and age. The results of experiments in which lysates with 'low' and 'high' activities were mixed gave no indication that individual variations in RBC HNMT activities were due to the effects of endogenous enzyme inhibitors or activators. RBC HNMT activities measured in blood samples from 17 individual subjects four times over 6 wk were quite constant in each subject, with an average coefficient of variation of 6.2%. The availability of this assay will make it possible to test the hypothesis that individual variations in determined variations in human RBC COMT activity are directly correlated with variations in COMT activity in other human tissues [8,9]. Those variations are also significantly correlated with large individual differences in the O-methylation by COMT of orally administered catechol drugs such as l-dopa and methyldopa [32,33]. RBC TPMT activity in man is also under genetic control , and the properties and levels of activity of RBC TPMT reflect those of the enzyme in other human tissues and cells [10,11]. The results of our experiments with RBC HNMT will now make it possible to determine whether individual variations in this human RBC methyltransferase may help us to understand and predict one factor responsible for individual variations in the NT-methylation of histamine and related compounds in man.
- Erythrocyte enzymes
- Histamine N-methyltransferase
ASJC Scopus subject areas
- Clinical Biochemistry
- Biochemistry, medical