NAD+ and NADP+ as Prosthetic Groups for Enzymes


NAD(P)+ (nicotinamide–adenine dinucleotide (phosphate)) serves as a coenzyme for pyridine nucleotide‐dependent dehydrogenase catalysed redox reactions. When the coenzyme behaves as a nondissociable prosthetic group it can function to promote aldehyde dismutation with nicotinoproteins as well as regular alcohol dehydrogenases. The tightly bound coenzyme can also be used to conduct reversible redox reactions to catalyse epimerization of sugar hydroxyls and to trigger elimination reactions; for example, nucleotidyl diphosphohexose‐4,6‐dehydratases, ornithine cyclodeaminase and S‐adenosyl‐l‐homocysteine hydrolase. Finally in urocanase the nicotinamide ring can participate directly as a Lewis acid to catalyse a complex rearrangement reaction. An important feature of these reactions is that because the coenzyme does not dissociate that they are independent of the NAD+/NADH (reduced form of nicotinamide–adenine dinucleotide) ratio of the cell.

Key Concepts:

  • In the standard spectrophotometric assay of dehydrogenases coenzyme release is rate limiting, but when the coenzyme cannot or does not dissociate then chemistry becomes limiting and reaction rates can be much faster when substrates are present for both forward and reverse reactions.

  • Dismutation of aldehydes and reactions catalysed by nicotinoproteins can occur without any change in the concentration of exogenous NADH, that is, no spectrophotometric change hence they are difficult to observe and assay.

  • For enzymes with tightly bound coenzyme, their reactions are not directly influenced by changes in the ratio of NAD+/NADH in the cell.

  • With the exception of nicotinoproteins, enzymes with nondissociable NAD+ can be inactivated by its reduction to NADH in the absence of the normal substrate, that is, there is no independent means of exchanging out the bound NADH or oxidizing it.

  • In spite of the chiral nature of their active sites epimerases catalyse an overall nonstereospecific reaction. Although the hydride transfer reaction retains its exquisite stereochemical fidelity the binding of the substrate has greatly decreased stringency allowing reactions with either epimer, hence eventually leading to the thermodynamically favoured mixture.

  • In addition to redox reactions the catalytic machinery of dehydrogenases can promote a wide range of other chemistry including dehydrations and rearrangements provided the substrate can be properly activated and bound in the appropriate juxtaposition to the coenzyme.

  • The pyridinium ring of NAD+ can serve as a Lewis acid for reactions where protonation does not provide the correct pathway.

Keywords: dismutation; NAD; epimerization; dehydrogenase; nicotinoproteins

Figure 1.

Alcohol dehydrogenase‐catalysed dismutation of aldehydes.

Figure 2.

Nicotinoprotein‐catalysed oxidation of alcohols.

Figure 3.

Mechanism of nucleotidyl diphosphohexose epimerases.

Figure 4.

Mechanism of S‐adenosyl‐l‐homocysteine hydrolase.

Figure 5.

Mechanism of nucleotidyl diphosphohexose 4,6‐dehydratases.

Figure 6.

Mechanism of ornithine cyclodeaminase.

Figure 7.

Mechanism of urocanase. (Mechanism has been redrawn to illustrate the involvement of active site amino acid residues.)



Abeles RH and Lee HA Jr (1960) The dismutation of formaldehyde by liver alcohol dehydrogenase. Journal of Biological Chemistry 235: 1499–1503.

Abeles RH, Fish S and Lapinskas B (1982) S‐adenosylhomocysteinase: mechanism of inactivation by 2′‐deoxyadenosine and interaction with other nucleosides. Biochemistry 21(22): 5557–5562.

Daussmann T, Aivasidis A and Wandrey C (1997) Purification and characterization of an alcohol:N,N‐dimethyl‐4‐nitrosoaniline oxidoreductase from the methanogen Methanosarcina barkeri DSM 804 strain Fusaro. European Journal of Biochemistry 248(3): 889–896.

De Clercq E (2005) John Montgomery's legacy: carbocyclic adenosine analogues as SAH hydrolase inhibitors with broad‐spectrum antiviral activity. Nucleosides, Nucleotides & Nucleic Acids 24(10–12): 1395–1415.

Frey PA (1987) Complex pyridine nucleotide‐dependent transformations. In: Dolphin D, Poulson R and Avramovic O (eds) Pyridine Nudeotide Coenzyrnes: Chemical, Biochemical and Medical Aspects, pp. 461–511. New York: Wiley.

Gerlinger E, Hull WE and Retey J (1981) Mechanistic study of the urocanase reaction using deuterated substrates and 1H‐NMR spectroscopy. European Journal of Biochemistry 117(3): 629–634.

Goodman JL, Wang S, Alam S et al. (2004) Ornithine cyclodeaminase: structure, mechanism of action, and implications for the mu‐crystallin family. Biochemistry 43(44): 13883–13891.

Henehan GT and Oppenheimer NJ (1993) Horse liver alcohol dehydrogenase‐catalyzed oxidation of aldehydes: dismutation precedes net production of reduced nicotinamide adenine dinucleotide. Biochemistry 32(3): 735–738.

Henehan GT, Chang SH and Oppenheimer NJ (1995) Aldehyde dehydrogenase activity of Drosophila melanogaster alcohol dehydrogenase: burst kinetics at high pH and aldehyde dismutase activity at physiological pH. Biochemistry 34(38): 12294–12301.

Kato N, Yamagami T, Shimao M et al. (1986) Formaldehyde dismutase, a novel NAD‐binding oxidoreductase from Pseudomonas putida F61. European Journal of Biochemistry 156(1): 59–64.

Kavanagh KL, Jornvall H, Persson B et al. (2008) Medium‐ and short‐chain dehydrogenase/reductase gene and protein families: the SDR superfamily: functional and structural diversity within a family of metabolic and regulatory enzymes. Cellular and Molecular Life Sciences 65(24): 3895–3906.

Kendal LP and Ramanathan AN (1951) Aldehyde mutase and the dismutation of formaldehyde. Biochemical Journal 49(4): lvii–lviii.

Kessler D, Retey J and Schulz GE (2004) Structure and action of urocanase. Journal of Molecular Biology 342(1): 183–194.

Kim RY, Gasser R and Wistow GJ (1992) mu‐crystallin is a mammalian homologue of Agrobacterium ornithine cyclodeaminase and is expressed in human retina. Proceedings of the National Academy of Sciences of the USA 89(19): 9292–9296.

Klepp J, Fallert‐Muller A, Grimm K et al. (1990) Mechanism of action of urocanase. Specific 13C‐labelling of the prosthetic NAD+ and revision of the structure of its adduct with imidazolylpropionate. European Journal of Biochemistry 192(3): 669–676.

Koivusalo M, Baumann M and Uotila L (1989) Evidence for the identity of glutathione‐dependent formaldehyde dehydrogenase and class III alcohol dehydrogenase. FEBS Letters 257(1): 105–109.

Liu S, Yuan CS and Borchardt RT (1996) Aristeromycin‐5′‐carboxaldehyde: a potent inhibitor of S‐adenosyl‐l‐homocysteine hydrolase. Journal of Medicinal Chemistry 39(12): 2347–2353.

Maxwell ES, De Robichon‐Szulmajster H and Kalckar HM (1958) Yeast uridine diphosphogalactose‐4‐epimerase, correlation between activity and fluorescence. Archives of Biochemistry and Biophysics 78(2): 407–415.

Miller A and Waelsch H (1957) The mechanism of urocanase action. Biochimica et Biophysica Acta 24(2): 447–448.

Montgomery JA, Clayton SJ, Thomas HJ et al. (1982) Carbocyclic analogue of 3‐deazaadenosine: a novel antiviral agent using S‐adenosylhomocysteine hydrolase as a pharmacological target. Journal of Medicinal Chemistry 25(6): 626–629.

Van Ophem PW, Van Beeumen J and Duine JA (1993) Nicotinoprotein [NAD(P)‐containing] alcohol/aldehyde oxidoreductases. Purification and characterization of a novel type from amycolatopsis methanolica. European Journal of Biochemistry 212(3): 819–826.

Oppenheimer NJ, Henehan GT, Huete‐Perez JA et al. (1997) P. putida formaldehyde dehydrogenase. An alcohol dehydrogenase masquerading as an aldehyde dehydrogenase. Advances in Experimental Medicine and Biology 414: 417–423.

Ozyurt AS and Selby TL (2008) Computational active site analysis of molecular pathways to improve functional classification of enzymes. Proteins 72(1): 184–196.

Palmer JL and Abeles RH (1979) The mechanism of action of S‐adenosylhomocysteinase. Journal of Biological Chemistry 254(4): 1217–1226.

Robins MJ, Wnuk SF, Yang X et al. (1998) Inactivation of S‐adenosyl‐l‐homocysteine hydrolase and antiviral activity with 5′,5′,6′,6′‐tetradehydro‐6′‐deoxy‐6′‐halohomoadenosine analogues (4′‐haloacetylene analogues derived from adenosine). Journal of Medicinal Chemistry 41(20): 3857–3864.

Sawaki S, Hattori N, Morikawa N et al. (1967) Oxidation and reduction of glyoxylate by lactate dehydrogenase. Journal of Vitaminology (Kyoto) 13(2): 93–97.

Takata Y, Yamada T, Huang Y et al. (2002) Catalytic mechanism of S‐adenosylhomocysteine hydrolase. Site‐directed mutagenesis of Asp‐130, Lys‐185, Asp‐189, and Asn‐190. Journal of Biological Chemistry 277(25): 22670–22676.

Thoden JB, Hegeman AD, Wesenberg G et al. (1997) Structural analysis of UDP‐sugar binding to UDP‐galactose 4‐epimerase from Escherichia coli. Biochemistry 36(21): 6294–6304.

Thoden JB, Wohlers TM, Fridovich‐Keil JL et al. (2001) Human UDP‐galactose 4‐epimerase. Accommodation of UDP‐N‐acetylglucosamine within the active site. Journal of Biological Chemistry 276(18): 15131–15136.

Tripp AE, Burdette DS, Zeikus JG et al. (1998) Mutation of Serine‐39 to threonine in thermostable secondary alcohol dehydrogenase from Thermoanaerobacter ethanolicus changes enantiospecificity. Journal of the American Chemical Society 120(21): 5137–5141.

Vogan EM, Bellamacina C, He X et al. (2004) Crystal structure at 1.8 A resolution of CDP‐d‐glucose 4,6‐dehydratase from Yersinia pseudotuberculosis. Biochemistry 43(11): 3057–3067.

Warren WA (1970) Catalysis of both oxidation and reduction of glyoxylate by pig heart lactate dehydrogenase isozyme 1. Journal of Biological Chemistry 245(7): 1675–1681.

Yang X, Yin D, Wnuk SF et al. (2000) Mechanisms of inactivation of human S‐adenosylhomocysteine hydrolase by 5′,5′,6′,6′‐tetradehydro‐6′‐deoxy‐6′‐halohomoadenosines. Biochemistry 39(49): 15234–15241.

Further Reading

Allard ST, Giraud MF and Naismith JH (2001) Epimerases: structure, function and mechanism. Cellular and Molecular Life Sciences 58(11): 1650–1665.

Contact Editor close
Submit a note to the editor about this article by filling in the form below.

* Required Field

How to Cite close
Oppenheimer, Norman J(Apr 2010) NAD+ and NADP+ as Prosthetic Groups for Enzymes. In: eLS. John Wiley & Sons Ltd, Chichester. [doi: 10.1002/9780470015902.a0000637.pub2]