Metabolic Profiling of Alternative NAD Biosynthetic Routes in Mouse Tissues

Abstract

NAD plays essential redox and non-redox roles in cell biology. In mammals, its de novo and recycling biosynthetic pathways encompass two independent branches, the “amidated” and “deamidated” routes. Here we focused on the indispensable enzymes gating these two routes, i.e. nicotinamide mononucleotide adenylyltransferase (NMNAT), which in mammals comprises three distinct isozymes, and NAD synthetase (NADS). First, we measured the in vitro activity of the enzymes, and the levels of all their substrates and products in a number of tissues from the C57BL/6 mouse. Second, from these data, we derived in vivo estimates of enzymes'rates and quantitative contributions to NAD homeostasis. The NMNAT activity, mainly represented by nuclear NMNAT1, appears to be high and nonrate-limiting in all examined tissues, except in blood. The NADS activity, however, appears rate-limiting in lung and skeletal muscle, where its undetectable levels parallel a relative accumulation of the enzyme's substrate NaAD (nicotinic acid adenine dinucleotide). In all tissues, the amidated NAD route was predominant, displaying highest rates in liver and kidney, and lowest in blood. In contrast, the minor deamidated route showed higher relative proportions in blood and small intestine, and higher absolute values in liver and small intestine. Such results provide the first comprehensive picture of the balance of the two alternative NAD biosynthetic routes in different mammalian tissues under physiological conditions. This fills a gap in the current knowledge of NAD biosynthesis, and provides a crucial information for the study of NAD metabolism and its role in disease.

Replicatively senescent human fibroblasts reveal a distinct intracellular metabolic profile with alterations in NAD+ and nicotinamide metabolism

“…Nicotinate can be converted to NAD+ from nicotinate mononucleotide involving the enzymes NMNAT and NADS via the deamidated route31. However, extracellular nicotinate was specifically depleted in PEsen cells as reported previously23 and intracellular nicotinate was undetectable, nicotinate mononucleotide, nicotinate riboside and deamido NAD+ as well as NMNAT 1 and 3 were all undetectable in this study.…”

Biological Activities of the Essential Oil from Erigeron floribundus

“…A preliminary single point inhibition test at the concentration of 80 µg/mL indicated a~60% inhibition with no effect on the ubiquitous mammalian isoform NMNAT1 [28]. Motivated by these results and by an observed broad-spectrum antimicrobial activity based on the diffusion IZD and MIC methods (Table 3), we performed a dose-response inhibition using an oil concentration range of 5-160 µg/mL.…”

Physiological and pathophysiological roles of NAMPT and NAD metabolism

“…NAM, rather than nicotinic acid, is thought to be the predominant NAD precursor in mammals. [8][9][10] NAM is also a product of deacetylation and ADP-ribosylation reactions, which are catalysed by NAD-dependent enzymes. [11][12][13] NAMPT activity generates NMN from NAM and 5'-phosphoribosyl-1-pyrophosphate (PRPP), thereby catalysing the rate-limiting step in the mammalian NAD salvage pathway from NAM.…”