NAD+ is an essential cofactor of metabolic reactions and also acts as a substrate for important enzymes such as Sirtuins. Sirtuins are NAD+ dependent enzymes fast emerging as key biological players involved in energy metabolism and mitochondrial functioning, both factors closely implicated in the aging process as well as the development of age-related health conditions. In recent years, a better understanding of NAD+ metabolism and their role in Sirtuins activation has sparked an interest in developing strategies to enhance NAD+ levels to manage a range of age-related diseases – including diabetes, obesity, Alzheimer’s, Parkinson’s and even cancer.
What is NAD?
NAD – Nicotinamide adenine dinucleotide (NAD) is a coenzyme found in all living cells. This ubiquitous molecule is an integral part of redox and metabolic reactions in the body, carrying electrons from one reaction site to another. It exists in two forms; oxidized (NAD+) and reduced (NADH) states.
NAD+ (oxidized form of NAD) is an oxidizing agent and gains electrons during the oxidation of organic molecules, such as carbohydrates, proteins and fatty acids. It is reduced to NADH in the process. NAD+ is a cornerstone of energy metabolism within the mitochondria, commonly referred as the powerhouse of the cells. Emerging studies show that NAD+ plays key roles in longevity, anti-aging effects mediated by calorie-restriction and many diseases that are associated with age.
NADH (reduced form of NAD) is a reducing agent. Reducing agents are substances that donate electrons and which is oxidized in the process. NADH transports electrons to the mitochondria, where the cells harvest the energy stored in these high energy electrons to create ATP. NADH is cycled back in the body to NAD+ through NAD salvage pathways.
How NAD+ works?
NAD+ has two overarching roles in the body:
- NAD+ serves as an extremely important cofactor in redox and metabolic reactions, where it shuttles back and forth between the oxidized (NAD+) and reduced (NADH) states. For example, NAD+ is reduced to NADH during one of the steps in the glycolysis cycle, a metabolic pathway that involves the breakdown of glucose to release energy.
NAD+ is also converted to NADH during the oxidation of fatty acids, carbohydrates and amino acids in mitochondria, where NADH donates the reduced electrons to produce ATP molecules via oxidative phosphorylation process. ATP, as we know, is referred to as energy currency and is required for cellular metabolism. In such reactions, NAD+ is not consumed.
- NAD+ is also used as a substrate for enzymes such as Sirtuins, ADP-ribose transferases, poly(ADP-ribose) polymerases (PARPs) and cADP-ribose synthases. In such reactions, NAD+ is completely consumed and not recycled.
(Substrates are molecules that enzymes act upon. These specific molecules bind onto the active site of the enzymes resulting in enzyme-substrate complex. The chemical reactions result in the formation of a new product that ultimately separates from the enzyme. While the enzyme goes on to catalyze or speed up other bio-chemical reactions, substrate is transformed into another molecule or product.)
NAD+ and Sirtuins
Sirtuins are NAD+ dependent enzymes, which play key roles in promoting health, longevity, stress resistance, double stranded DNA repair and regulating metabolic pathways. The Sirtuin family consists of seven enzymes in mammals, SIRT1–7, each with its own unique enzymatic actions. Specifically, SIRT1 and SIRT3 are researched for their anti-aging effects and are believed to be closely related to longevity by turning off certain genes that speed up aging.
NAD+ is an essential cofactor of the Sirtuin family of enzymes. Basically, these key enzymes require NAD+ to remove acetyl groups from proteins responsible for regulating stress, energy metabolism, DNA repair and cell survival pathways. Sirtuins are important actors in optimizing mitochondrial functions too. Sirtuins improve metabolic efficiency as well as up-regulating the mitochondrial response to oxidative stress. Besides the Sirtuins family, two more known classes of enzymes compete for the NAD+ pool in the nucleus; namely Poly-ADP-ribose Polymerases (PARPs) and CD38. All of these classes of proteins require NAD+ as a substrate to perform important functions required to maintain optimal cell health.
NAD+ levels in the mitochondria reflect the energy status of the cell. You can say Sirtuins act as metabolic sensors that sense changes in the NAD+ levels within the cells and use this information to help cells modulate metabolic pathways accordingly . Such adaptations are important to meet the energy requirements at hand. Sirtuin, thus, play crucial roles in cellular response to nutritional and environmental stimuli, such as fasting, oxidative stress and DNA damage.
NAD+ dependent activation of Sirtuins is believed to be valuable for metabolism related conditions such as type 2 diabetes and obesity and also for neurodegenerative diseases such as Alzheimer’s and Parkinson’s disease. Sirtuins work by stimulating the activity of mitochondria and of mitochondrial proteins involved in these conditions. Long story short, Sirtuins influence multiple cellular pathways involved in increasing the life span and improving overall health.
NAD+ and Mitochondrial Health
Mitochondria generate chemical energy from the available nutrients to carry out important biological functions implicated in cellular growth and survival. These double-membrane bound tiny organelles play integral roles in energy production, metabolism and intracellular signalling.
Mitochondria are highly dynamic in their ability to alter their number, function and structure in response to environmental stressors such as diet and nutrient availability, exercise, temperature and hormones. Basically, the number, function and capacity of mitochondria continually modulates with the fluctuating energy demands of cells and tissues.
One of the main functions of mitochondria is to generate ATP through the process of oxidative phosphorylation. Oxidation of glucose, amino acids and fatty acids releases high-energy electrons and their energy is used to pump protons from the mitochondrial matrix to the inner membrane zone, ultimately driving the synthesis of ATP through a series of complicated reactions and mechanisms. Free radicals or reactive oxygen species (ROS) are formed as a side-product of this energy creation process. These free radicals react with fragile biological molecules such as lipids, proteins and mitochondrial DNA (mtDNA), causing oxidative stress. The increased oxidative stress is cited as one of the main causes of premature cellular aging (also referred to as the mitochondrial theory of aging).
Properly functioning mitochondria are crucial to maintaining metabolic balance and triggering suitable cellular responses to stressors. It is hardly any surprise that mitochondrial dysfunction is linked with aging and age-associated diseases such as type 2 diabetes, Alzheimer’s, cancer and cardiovascular disease. So, how does NAD+ fit in here? The NAD+ /Sirtuin axis is known to play a predominant role in optimizing mitochondrial function.
NAD+ as a cofactor in redox reactions
We understand how NAD+ plays a critical role in transferring energy – derived from the oxidation of organic fuel – to the mitochondria to generate ATP. In the absence of sufficient NAD+, the energy transfer takes a hit. With low levels of NAD+, mitochondrial fitness deteriorates and the number and concentration of the mitochondria are reduced – resulting in mitochondrial inefficiency and even dysfunction. This incompetence manifests in the form of decline in ATP generation (needed for growth and life-sustenance) and excessive production of free radicals (triggering mutations in the mitochondrial DNA). This clearly hampers a cell’s ability to perform its functions such as reproduction, growth, repair and maintenance – contributing to the aging process and the accompanying pathologies.
NAD+ as a substrate for Sirtuins
NAD+ stimulates the activity of Sirtuins, which helps in forming new mitochondria and keep mitochondria running smoothly. Sirtuins also help the mitochondria to develop resistance against oxidative stress by increasing certain anti-oxidant pathways and enabling repair of damaged DNA through deacetylation or modification of proteins involved in the repair. Again, Sirtuins are dependent on NAD+ to carry out their protein deacetylation activities.
Decline in NAD+ levels
NAD+ levels decline as we age. Overeating, a high-fat diet and sedentary lifestyles are also known to diminish the levels of NAD+.
NAD+ facilitates the communication within the cell between the nucleus and its mitochondria, thus maintaining mitochondrial efficiency. With diminishing NAD+ levels, this communication is interrupted and aging is accelerated . Dr. David Sinclair, Harvard Medical School Professor of Genetics, says this communication network is “like a married couple—when they are young, they communicate well, but over time, living in close quarters for many years, communication breaks down. And just like with couples, restoring communication solves the problem.”
NAD+ Health Benefits
We just read how NAD+ serves as a vital cofactor as well as substrate that can regulate metabolism, activate the Sirtuins family of enzymes and maintain mitochondrial health. Falling levels of this important metabolite and the subsequent impact on the activity of NAD+ dependent Sirtuins are considered as one of the main reasons behind metabolic disturbance and mitochondria dysfunction – resulting in the development and progression of premature aging and the age-related diseases.
“NAD+ levels decline during the aging process and may be an Achilles’ heel, causing defects in nuclear and mitochondrial functions and resulting in many age-associated pathologies. Restoring NAD+ by supplementing NAD+ intermediates can dramatically ameliorate these age-associated functional defects, counteracting many diseases of aging, including neurodegenerative diseases. Thus, the combination of Sirtuin activation and NAD+ intermediate supplementation may be an effective antiaging intervention, providing hope to aging societies worldwide.” 
Benefits of maintaining NAD+ levels in the body
Maintaining optimum levels of NAD+ in the body is important for maintaining overall health and preventing, and even reversing, a range of diseases. NAD+ is critical for:
- Improved energy levels and vitality
- Healthy metabolism
- Longevity; Control of inflammatory processes and premature aging factors
- Overall healthy and disease free life
- Healthy cognitive functions
Studies show that beneficial effects of NAD+ may extend to prevent, reverse or treat the following conditions:
- Neuro-degenerative diseases; Alzheimer’s and Parkinson’s
- Metabolic disorders; Obesity, insulin resistance and type 2 diabetes
- Cardiovascular diseases
- Inherited mitochondrial diseases
- Cantó C, Auwerx J. Targeting Sirtuin 1 to improve metabolism: all you need is NAD(+)? Pharmacological Reviews 2012.
- Gomes et al. Declining NAD+ Induces a Pseudohypoxic State Disrupting Nuclear-Mitochondrial Communication during Aging. Cell. 2013.
- Imai et al. NAD+ and Sirtuins in Aging and Disease. Trends in Cell Biology. 2014.