52 by: Trevor Lacey

Student – Trevor Lacey

Enzyme – Nicotinate Dehydrogenase

E.C. # –  1.17.1.5

This enzyme is found in the obligate anaerobe Eubacteria barkeri, formerly Clostridium barkeri, as a cytoplasmic enzyme. There is an analogue in Pseudomonas that is bound to
the cell membrane and uses oxygen as the terminal electron accepter with an electron transfer chain. This enzyme performs the first step in the fermentation pathway for nicotinic acid, also known as niacin or vitamin B3. The enzyme oxidizes the substrate, replacing a hydrogen on the pyridine ring with a hydroxyl group obtained from water.  The mechanism begins with substrate binding. An arginine residue in the active site forms hydrogen bonds with the substrate’s carboxyl group. At cellular pH, the arginine side chain should be protonated and the substrate carboxyl is deprotonated. Another hydrogen bond forms between the nitrogen atom and a tyrosine residue. The hydroxyl group on molybdenum attacks C6, and a basic glutamic acid residue accepts the proton. At the same time, the C6 hydrogen leaves as a hydride ion, transferring both electrons to selenium (Wagener, Pierik, Ibdah, Hille, & Dobbek, 2009). At this point I couldn’t find further information, but we know the enzyme must be regenerated, the substrate must leave with a hydroxyl from H2O, and the hydride makes its way to NADP+. I modeled a water molecule as an acid that donates a proton to the Mo-O-substrate bond before attacking it as a nucleophile. A review of other molybdenzymes is helpful. Xanthine oxidase, which seems similar, shuttles the electron pair through [2Fe-2S] clusters to FAD cofactor, and ultimately to the terminal electron receptor (O2 or NAD+) (Adamus, Ruszczynska, & Wyczalkowska- Tomasik, 2024). I assume it must be a similar situation in NDH, since it shares many of those cofactors and the NADP+ is distant from the active site in 3d models. It is a dimer of heterotretramers, each 160kDa tetramer having independent activity. The enzyme’s specificity lies in the substrate’s aromatic N and a carboxy group meta to it. The selenium in this enzyme is labile. That is, it is not incorporated in a cysteine residue like other selenol enzymes.

References
Adamus, J., Ruszczynska, A., & Wyczalkowska-Tomasik, A. (2024). Molybdenum’s Role as an Essential Element in Enzymes Catabolizing Redox Reactions: A Review. Biomolecules,
869.
Holcenberg, J., & Stadtman, E. (1969). Nicotinic Acid Metabolism: Purification and Propteries of a Nicotinic Acid Hydroxylase. The Journal of Biological Chemistry, 244(5), 1194-
1203.
Information on EC 1.17.1.5 – nicotinate dehydrogenase. (n.d.). Retrieved from BRENDA: https://www.brenda-enzymes.org/enzyme.php?ecno=1.17.1.5
Wagener, N., Pierik, A., Hille, R., & Dobbek, H. (2009). Crystal Structure of Nicotinate Dehydrogenase. doi:https://doi.org/10.2210/pdb3HRD/pdb
Wagener, N., Pierik, A., Ibdah, A., Hille, R., & Dobbek, H. (2009). The Mo-Se active site of nicotinate dehydrogenase. Proceedings of the National Academy of Sciences of the United
States of America, 106(27), 11055-11060. doi:https://doi.org/10.1073/pnas.0902210106