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NAD+ And The Science of Metabolism & Healthspan

Summary

  1. NAD+ is crucial to proper cell function.
  2. When cells are under acute stress from a disease or even simple sun exposure, there is NAD+ breakdown.
  3. In multiple studies, NIAGEN® produced by ChromaDex, Inc. and tested in laboratories worldwide, has been shown to boost levels of NAD+.

NAD & human health: key findings over time

Why is NAD+ so important? The question is simple, but the answer stretches back over a century. To get the scientific perspective, ProHealthspan went to the world’s foremost authority on NAD+ metabolism: Charles Brenner, Ph.D.

In 2004, Dr. Brenner discovered that nicotinamide riboside (NR), a natural product found in milk, is a vitamin precursor of coenzyme NAD+ (nicotinamide adenine dinucleotide, also known as simply NAD—pronounced “en – A – dee”). This discovery ignited the science and research around nutritional supplements, NAD+ and aging.

While the power and benefits of NAD+ had been researched for years, Dr. Brenner’s breakthrough revealed a new and more efficient way to synthesize NAD+. Thanks to Dr. Brenner’s landmark work, today we know not only that NAD+ is crucial for metabolism, but also that NR is a safe and effective means to increase NAD+ levels in people.

The first and only commercially available form of NR is called NIAGEN®. Only one company—ChromaDex, Inc.—holds the worldwide patents on NIAGEN®. ProHealthspan has partnered with ChromaDex to market the NIAGEN® supplement called TRU NIAGEN®.

NAD+ is required for all bioenergetic processes of breaking down food and generating ATP—the crucial chemical in human metabolism that gives cells their ability to do work. For the heart to beat, for blood to flow, for our eyes to see, and our brains to conceive of ideas—we have to convert biological fuels into metabolites like ATP using the NAD+ coenzyme. NAD+ is also required for the function of enzymes that assist in DNA repair, regulate gene expression, and mediate anti-aging activities of cells.

Content produced in collaboration with Dr. Charles Brenner, the Roy J. Carver Chair & Head of Biochemistry, Professor of Internal Medicine, and founding Co-Director of the Obesity Research and Education Initiative at the University of Iowa.

Discovery of NAD+ as cozymase

At the turn of the 20th century, work on the metabolism of glucose was foundational for the discovery and elucidation of the NAD+ co-enzyme. First, Dr. Eduard Büchner—the inventor of the sintered glass Büchner funnel—showed that you don’t need living yeast cells but rather a yeast extract to ferment glucose. Second, Dr. Arthur Harden proved that the glucose-fermenting activity was due to a protein fraction he called zymase and a metabolite fraction he called cozymase. We now call zymase “enzymes” and cozymase “NAD<sup>+</sup>.”                                                                                                

The word “enzyme” literally means “in yeast” but enzymes are as critical to human metabolism as they are to yeast metabolism. Enzymes metabolize the fuels in our diets (carbohydrates, proteins and fats) but they can’t do this without the NAD+ coenzyme.

Structure of NAD+ and two vitamins

The chemical structure of NAD+ was determined independently by Drs. Hans von Euler and Otto Warburg in 1936. They showed that NAD+ contained a ring-shaped molecule called nicotinamide linked to ribose, two phosphates, and adenosine. In the next two years, two vitamin precursors of NAD+ were discovered by Dr. Conrad Elvehjem. These compounds, nicotinamide (sometimes called niacinamide) and nicotinic acid (sometimes called niacin or nicotinate) prevent a nutritional deficiency termed pellagra that once affected more than 1 million people in the American south. People with pellagra don’t have enough NAD+ precursors in their diet. Before nicotinamide and nicotinic acid were discovered as vitamins, one of the ways that pellagra was prevented was with a pint of milk.

Pathways to NAD+

In the 1940s, the amino acid tryptophan was shown to be an NAD+ precursor. It was calculated that it takes 60 grams of tryptophan to be the equal of 1 gram of nicotinic acid or nicotinamide vitamins.

Based on this knowledge, nicotinic acid and nicotinamide became widely supplemented into foods and nicotinic acid, in particular, gained use as a cholesterol drug. However, high doses of nicotinic acid cause a side effect called the niacin flush in which the skin becomes red and hot. With niacin supplementation, pellagra became a rare condition. Recommended daily allowances of niacin were established but it’s not clear whether those levels are truly optimal for human health.

Sirtuins break NAD+ in two

If you have been following this story, you know that nicotinamide is a piece of NAD+ (von Euler and Warburg taught us that) and nicotinamide is also a vitamin precursor of NAD+ (Elvehjem taught us that).

NAD+ is a coenzyme, which means it is a catalyst for metabolic reactions—when fuels are converted to ATP, the NAD+ is not broken down but can be used over and over again.

However, it became apparent that NAD+ is broken down inside of every cell. In fact, it started becoming clear at the turn of the 21th century that NAD+ is broken down when DNA is damaged, when cells turn on calcium signals, and when cells change their gene expression programs, switching from one metabolic state to another. These new roles for NAD+ attracted more scientists to investigate NAD-dependent processes of relevance to human health.

Aging is a co-factor in almost all diseases, which is to say that the older we get, the greater is our risk for lots of different maladies and the harder it is for us to recover. Just as Büchner and Harden used yeast as a model for glucose metabolism, Dr. Leonard Guarente set out to determine how a yeast cell ages as a model for how animals age.

Guarente knew there was a connection between gene regulation and aging because extra copies of the yeast SIR2 gene keep yeast cells young. SIR2 stands for silent information regulator 2 and it encodes an enzyme that is responsible for turning genes off. The worm SIR2 gene can also extend the life of worms. Seven different forms of this gene—sirtuins—are found in mice and in people. 

Guarente showed that the effects of sirtuins in gene regulation and opposing aging depend on NAD+. Interestingly, sirtuins don’t use NAD+ as a coenzyme that can be recycled over and over again. Sirtuins break NAD+ into two pieces: nicotinamide plus the rest of the molecule, which is called ADPribose. Similarly, the enzymes that respond to DNA damaged and that produce calcium-mobilizing signals break NAD+ into nicotinamide plus ADPribose products. Brenner coined the term NAD+-consuming enzymes to emphasize that enzymes such as sirtuins break NAD+ into two pieces. NAD+-consuming enzymes create the requirement that cells have to constantly replenish NAD+, lest they become NAD+-deficient, which ultimately leads to pellagra.

Nicotinamide riboside as the third vitamin precursor of NAD+

By 2004, NAD+ biology had entered an exciting phase. Many groups began exploring sirtuin activities in mouse and human systems. The genes required to convert tryptophan, nicotinamide and nicotinic acid to NAD+ were known, so people started to manipulate them to look at their influence on gene expression and aging. The Brenner group decided to question the received wisdom of the past and designed an experiment to determine whether there might be a third NAD+ precursor vitamin. Sure enough, Brenner discovered nicotinamide riboside (NR) as an unanticipated vitamin that is converted to NAD+ by virtue of nicotinamide riboside kinases. Brenner further discovered that nicotinamide riboside kinase genes are found in people and that milk is a source of NR. He went on to show that NR supplementation of yeast cells increases SIR2 activity and extends lifespan.

NAD+ precursor vitamins are not identical

There are distinct genes and enzymes that convert tryptophan, nicotinic acid, nicotinamide and NR to NAD+. Every tissue can make NAD+ from nicotinamide and from NR but not every tissue can convert tryptophan or nicotinic acid to NAD+. Cells that are damaged—including damaged nerves and failing heart—turn on nicotinamide riboside kinase 2, which tells us that NR is prized by cells when they are stressed out. High dose nicotinic acid produces the flush reaction and high dose nicotinamide inhibits sirtuins but NR does not have either of these limitations. Remarkably, NAD+ metabolism declines in aging and is disturbed in a number of diseases. NR is being tested in laboratory and clinical trials around the world to protect health and healthspan.

In more than 35 additional publications and issued patents on NAD+, Dr. Brenner identified previously-unknown steps in NAD+ metabolism, showed that NR increases sirtuin activity and lifespan in model systems, and developed the technology to quantify the NAD+ metabolome in any cell system, including milk and blood.

Summary

  1. NAD+ is crucial to proper cell function.
  2. When cells are under acute stress from a disease or even simple sun exposure, there is NAD+ breakdown.
  3. In multiple studies, NIAGEN® produced by ChromaDex, Inc. and tested in laboratories worldwide, has been shown to boost levels of NAD+.

    It’s important to note that NR has been tested in multiple animal models of aging, brain function and resistance to the ravages of high-fat diets. Of course, animals are not humans. In 2015, the first human clinical studies were completed and the results showed that NR boosts levels of NAD+ in healthy human volunteers. The study also revealed no safety issues with NR, which is consistent with the safety results demonstrated in animal studies.*

    Of Note

    See below for the significant papers published on NAD+, with links to their respective pdf reprints. Especially important are numbers 4, 8, 9, 10, 12, 17, 34 and 35.

    1. P. Bieganowski, H.C. Pace & C. Brenner, "Eukaryotic NAD+ Synthetase Qns1 Contains an Essential, Obligate Intramolecular Thiol Glutamine Amidotransferase Domain Related to Nitrilase," J Biol Chem, v. 278, pp. 33049-33055 (2003).

    2. P. Bieganowski & C. Brenner, "The Reported Human NADsyn2 is Ammonia-Dependent NAD+ synthetase from a Pseudomonad," J Biol Chem, v. 278, pp. 33056-33059 (2003). 

    3. D.A. Kwasnicka, A. Krakowiak, C. Thacker, C. Brenner & S.R. Vincent, " Coordinate Expression of NADPH-Dependent Flavin Reductase, FRE-1, and Hint-Related 7meGMP-Directed Hydrolase, DCS-1," J Biol Chem, v. 278, pp. 39051-39058 (2003). 

    4. P. Bieganowski & C. Brenner, "Discoveries of Nicotinamide Riboside as a Nutrient and Conserved NRK Genes Establish a Preiss-Handler Independent Route to NAD+ in Fungi and Humans," Cell, v. 117, pp. 495-502 (2004). 

    5. C. Brenner, "Evolution of NAD+ Biosynthetic Enzymes," Structure, v. 13, pp. 1239-1240 (2005). 

    6. P. Bieganowski, H.F. Seidle, M. Wojcik & C. Brenner, "Synthetic Lethal and Biochemical Analyses of NAD+ and NADH Kinases in Saccharomyces cerevisiae Establish Separation of Cellular Functions," J Biol Chem, v. 281, pp. 22439-22445 (2006). 

    7. M. Wojcik, H.F. Seidle, P. Bieganowski & C. Brenner, "Glutamine-Dependent NAD+ synthetase: How a Two-Domain, Three-Substrate Enzyme Avoids Waste," J Biol Chem, v. 281, pp. 33395-33402 (2006). 

    8. P. Belenky, K.L. Bogan & C. Brenner, "NAD+ Metabolism in Health and Disease," Trends in Biochemical Sciences, v. 32, pp. 12-19 (2007). 

    9. P. Belenky, F.G. Racette, K.L. Bogan, J.M. McClure, J.S. Smith & C. Brenner, "Nicotinamide Riboside Promotes Sir2 Silencing and Extends Lifespan via Nrk and Urh1/Pnp1/Meu1 Pathways to NAD+," Cell, v. 129, pp. 473-484 (2007). 

    10. W. Tempel, W.M. Rabeh, K.L. Bogan, P. Belenky, M. Wojcik, H.F. Seidle, L. Nedyalkova, T. Yang, A.A. Sauve, H.-W. Park & C. Brenner, "Nicotinamide Riboside Kinase Structures Reveal New Pathways to NAD+," PLoS Biology, v. 5, issue 10, e263 (2007). 

    11. P.A. Belenky, T.G. Mogu & C. Brenner, "S. cerevisiae YOR071C Encodes the High Affinity Nicotinamide Riboside Transporter, Nrt1," J Biol Chem, v. 283, pp. 8075-8079 (2008). 

    12. K.L. Bogan & C. Brenner, "Nicotinic Acid, Nicotinamide, and Nicotinamide Riboside: A Molecular Evaluation of NAD+ Precursor Vitamins in Human Nutrition," Ann Review Nutrition, v. 28, pp. 115-130 (2008). 

    13. P. Belenky, K.C. Christensen, F. Gazzaniga, A.A. Pletnev & C. Brenner, "Nicotinamide Riboside and Nicotinic Acid Riboside Salvage in Fungi and Mammals: Quantitative Basis for Urh1 and Purine Nucleoside Phosphorylase Function in NAD+ Metabolism," J Biol Chem, v. 284, pp. 158-164 (2009). 

    14. P. Bieganowski & C. Brenner, "Nicotinamide Riboside Kinase Compositions and Methods for Using the Same," Australian Patent 2005211773, issued June 1, 2009. 

    15. F. Gazzaniga, R. Stebbins, S. Z. Chang, M.A. McPeek & C.Brenner, "Microbial NAD Metabolism: Lessons from Comparative Genomics, "Microbiol Mol Biol Rev, v. 73, pp. 529-541 (2009). 

    16. K.L. Bogan, C. Evans, P. Belenky, P. Song, C.F. Burant, R.T. Kennedy & C. Brenner, " Identification of Isn1 and Sdt1 as Glucose and Vitamin-regulated NMN and NaMN 5'-nucleotidases Responsible for Production of Nicotinamide Riboside and Nicotinic Acid. Access here.Riboside," J Biol Chem, v. 284, pp. 34861-34869 (2009). 

    17. C. Evans, K.L. Bogan, P. Song, C.F. Burant, R.T. Kennedy & C. Brenner, "NAD+ Metabolite Levels as a Function of Vitamins and Calorie Restriction: Evidence for Different Mechanisms of Longevity," BMC Chem Biol, v. 10, 2 (2010). 

    18. K.L. Bogan & C. Brenner, "5'-Nucleotidases and their New Roles in NAD+ and Phosphate Metabolism," New Journal of Chemistry, v. 34, pp. 845-853 (2010). 

    19. C. Brenner. "On the Nonspecific Degradation of NAD+ to Nicotinamide Riboside," JBC, v. 286, p. le5 (2011). 

    20. P. Belenky, R. Stebbins, K.L. Bogan, C.R. Evans & C. Brenner, "Nrt1 and Tna1-Independent Export of NAD+ Precursor Vitamins Promotes NAD+ Homeostasis and Allows Engineering of Vitamin Production," PLoS ONE, v. 6, p. e19710 (2011). 

    21. C. Brenner, P. Belenky & K.L. Bogan, "Yeast Strain and Method for Using the Same to Produce Nicotinamide Riboside," US Patent 8,114,626, issued February 14, 2012. 

    22. R.R. Midtkandal, P. Redpath, S.A.J. Trammell, S.J.F. Macdonald, C. Brenner & M.E. Migaud, "Novel synthetic route to the C-nucleoside, 2-deoxy benzamide riboside," Bioorganic & Medicinal Chemistry Letters v. 22, pp. 5204-7 (2012). 

    23. C. Brenner, "Nicotinamide riboside kinase compositions and methods for using the same," US Patent8,197,807, issued June 12, 2012. 

    24. C. Brenner, "Nicotinamide riboside kinase compositions and methods for using the same," US Patent8,383,086, issued February 26, 2013. 

    25. K.L. Bogan & C. Brenner, "Biochemistry: Niacin/NAD(P)," Encyclopedia of Biological Chemistry, W.J. Lennarz & M.D. Lane, eds., v. 3, pp.172-178, (2013), Waltham, MA: Academic Press. 

    26. S.A.J. Trammell & C. Brenner, "Targeted, LCMC-Based Metabolomics for Quantitative Measurement of NAD+ Metabolites," Computational and Structural Biotechnology Journal, v. 4, e201301012 (2013). DOI: 10.5936/csbj.201301012. 

    27. S. Ghanta, R.E. Grossmann & C. Brenner, "Mitochondrial protein acetylation as a cell-intrinsic, evolutionary driver of fat storage: chemical and metabolic logic of acetyl-lysine modifications" Critical Rev Biochem & Mol Biol, v. 48, pp. 561-574 (2013). 

    28. S-C. Mei & C. Brenner, "NAD as a Genotype-Specific Drug Target" Chemistry & Biology, v. 20, pp. 1307-1308 (2013). 

    29. C. Brenner, "Metabolism: Targeting a fat-accumulation gene" Nature, v. 508, pp. 194-195 (2014). DOI: 10.1038/508194a. 


    30. S.-C. Mei & C. Brenner, "Quantification of Protein Copy Number in Yeast: the NAD+ Metabolome," PLoS Onev. 9, e106496 (2014). DOI: 10.1371/journal.pone.0106496. 

    31. C. Brenner, "Boosting NAD to Spare Hearing," Cell Metabolism, v. 21, pp.926-927 (2014). DOI: 10.1016/j.cmet.2014.11.015. 

    32. S.-C. Mei & C. Brenner, "Calorie Restriction-Mediated Replicative Lifespan Extension in Yeast Is Non-Cell Autonomous," PLoS Biology, v. 13, e1002048 (2015). 

    33. S.A.J. Trammell & C. Brenner, "NNMT: A Bad Actor in Fat Makes Good in Liver," Cell Metabolism, v. 22, pp. 200-201 (2015). DOI:10.1016/j.cmet.2015.07.017.

    34. S.A.J. Trammell, L. Yu, P. Redpath, M.E. Migaud & C. Brenner, "Nicotinamide Riboside Is a Major NAD+ Precursor Vitamin in Cow Milk," The Journal of Nutrition, v. 146, pp. 957-963 (2016). DOI: 10.3945/jn.116.230078. 

    35. S.A.J. Trammell, B.J.Weidemann, A.Chadda, M.S. Yorek, A. Holmes, L.J.Coppey, A. Obrosov, R.H. Kardon, M.A. Yorek & C. Brenner, "Nicotinamide Riboside Opposes Type 2 Diabetes and Neuropathy in Mice," Scientific Reports, v. 6, 26933 (2016). DOI: 10.1038/srep26933. 

     *These statements have not been evaluated by the Food and Drug Administration.  This product is not intended to diagnose, treat, cure, or prevent any disease.

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