Mitochondrial NAD kinase in health and disease

Abstract

Nicotinamide adenine dinucleotide phosphate (NADP), a co-enzyme and an electron carrier, plays crucial roles in numerous biological functions, including cellular metabolism and antioxidation. Because NADP is subcellular-membrane impermeable, eukaryotes compartmentalize NAD kinases (NADKs), the NADP biosynthetic enzymes. Mitochondria are fundamental organelles for energy production through oxidative phosphorylation. Ten years after the discovery of the mitochondrial NADK (known as MNADK or NADK2), a significant amount of knowledge has been obtained regarding its functions, mechanism of action, human biology, mouse models, crystal structures, and post-translation modifications. NADK2 phosphorylates NAD(H) to generate mitochondrial NADP(H). NADK2-deficient patients suffered from hyperlysinemia, elevated plasma C10:2-carnitine (due to the inactivity of relevant NADP-dependent enzymes), and neuronal development defects. Nadk2-deficient mice recapitulate key features of NADK2-deficient patients, including metabolic and neuronal abnormalities. Crystal structures of human NADK2 show a dimer, with the NADP⁺-binding site located at the dimer interface. NADK2 activity is highly regulated by post-translational modifications, including S188 phosphorylation, K76 and K304 acetylation, and C193 S-nitrosylation; mutations in each site affect NADK2 activity and function. In mice, hepatic Nadk2 functions as a major metabolic regulator upon increased energy demands by regulating sirtuin 3 activity and fatty acid oxidation. Hopefully, future research on NADK2 will not only elucidate its functional roles in health and disease but will also pave the way for novel therapeutics for both rare and common diseases, including NADK2 deficiency and metabolic syndrome.

Keywords: Antioxidation; MNADK; Mitochondria; NAD; NADK; NADK2; NADP.

Graphical abstract

Fig. 1

In mammalian cells, NADK2 phosphorylates NAD(H) to generate mitochondrial NADP(H). NAD(H) represents NAD⁺ and NADH. NADP(H) represents NADP⁺ and NADPH. NADK phosphorylates NAD⁺ to generate cytosolic NADP⁺.

Fig. 2

Hepatic NADK2 determines mitochondrial NADP(H) levels, maintains SIRT3 activity, and regulates fatty acid oxidation. In hepatocytes, NADK2 generates mitochondrial NADP(H), which is crucial to maintain the activities of NADP(H) dependent enzymes, such as AASS, DECR, and to regenerates antioxidant systems (left panel). Reduced level or activity of NADK2 decreases mitochondrial NADP(H) levels, activities of NADP(H) dependent enzymes, and SIRT3 activity, leading to increased levels of lysine, acetylation of SIRT3 targets and ROS. Overall, reduced NADK2 decreases FAO, leading to TG accumulation (right panel).

Fig. 3

Crystal structures of NADK2. A) alignment of NADK2 structures solved by independent studies. PDB codes: 7R4J and 7N29. B) NADK2 is organized in 2 domains, an N-terminal and a C-terminal domain. The extension, in green, located in the C-terminal domain, is critical for dimerization. C) surface plot of NADK2 structure, showing the critical extension and NADP⁺ binding. D) and E dimer of NADK2. F) NADP⁺ is located in dimer interface. PDB code: 7R4J. The protein structures are visualized by ChimeraX [45]. Also refer to the supplemental file for a video showing the 360° view. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 4

NADK2 post-translational modifications. NADK2 is phosphorylated at S188, acetylated at K76 and K304, and S-nitrosylated at C193. All the post-translational modifications have been characterized, and it has been demonstrated that mutations in each of the 4 sites affect NADK2 functions. A) all four sites are highly evolutionarily conserved. Locations of the post-translational modification sites in NADK2 B) monomer and C) dimer. The protein structures are visualized by ChimeraX [45]. Also refer to the supplemental file for a video showing the 360° view.