Nicotinamide N -methyltransferase (NNMT) is responsible for the N -methylation of nicotinamide to 1-methylnicotinamide. Our recent studies have demonstrated that NNMT regulates cellular processes fundamental to the correct functioning and survival of the cell. It has been proposed that NNMT may possess β-carboline N -methyltransferase activity, endogenously and exogenously-produced pyridine-containing compounds which, when N -methylated, are potent inhibitors of Complex I and have been proposed to have a role in the pathogenesis of Parkinson's disease. We have investigated the ability of recombinant NNMT to N -methylate norharman to 2- N -methylnorharman. In addition, we have investigated the toxicity of the β-carboline norharman, its precursor 1,2,3,4-tetrahydronorharman and its N -methylated metabolite 2- N -methylnorharman, using our in vitro SH-SY5Y NNMT expression model. Recombinant NNMT demonstrated norharman 2 N -methyltransferase activity, with a K m of 90 ± 20 µM, a k cat of 3 x 10-4 ± 2 x 10-5 s-1 and a specificity constant ( k cat/ K m) of 3 ± 1 s-1 M-1. 1,2,3,4-Tetrahydronorharman was the least toxic of all three compounds investigated, whereas norharman demonstrated the greatest, with no difference observed in terms of cell viability and cell death between NNMT-expressing and non-expressing cells. In NNMT-expressing cells, 2- N -methylnorharman increased cell viability and cellular ATP concentration in a dose-dependent manner after 72 and 120 h incubation, an effect which was not observed after 24 h incubation or in non-NNNT-expressing cells at any time point. Taken together, these results suggest that NNMT may be a detoxification pathway for β-carbolines such as norharman.
- enzyme kinetics
- ©2016 The Author(s)
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