Atomic Resolution Crystal Structure of NAD +Dependent Formate Dehydrogenase from Bacterium Moraxella sp. C-1

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Abstract


The crystal structure of the ternary complex of NAD--dependent formate dehydrogenase from the methylotrophic bacterium Moraxella sp. С -1 with the cofactor (NAD-) and the inhibitor (azide ion) was established at 1.1 A resolution. The complex mimics the structure of the transition state of the enzymatic reaction. The structure was refined with anisotropic displacements parameters for non-hydrogen atoms to a R factor of 13.4%. Most of the nitrogen, oxygen, and carbon atoms were distinguished based on the analysis of the temperature factors and electron density peaks, with the result that side-chain rotamers of histidine residues and most of asparagine and glutamine residues were unambiguously determined. A comparative analysis of the structure of the ternary complex determined at the atomic resolution and the structure of this complex at 1.95 A resolution was performed. In the atomic resolution structure, the covalent bonds in the nicotinamide group are somewhat changed in agreement with the results of quantum mechanical calculations, providing evidence that the cofactor acquires a bipolar form in the transition state of the enzymatic reaction.

Formate dehydrogenases catalyze the oxidation of formate into carbon dioxide and can be divided into several groups based on the quaternary structure, as well as on the presence of prosthetic groups and cofactors. The structurally simplest formate dehydrogenases are NAD--dependent formate dehydrogenases (FDH, EC 1.2.1.2), which oxidize formate coupled with reduction of the coenzyme NAD- into NADH [1]: НСОО- - NAD- ↔ СО2 ↑ - NADH. Formate dehydrogenases belong to a large superfamily of D-isomer specific 2-hydroxyacid dehydrogenases [2]. Formate dehydrogenases of this type contain no metal ions or prosthetic groups in the active sites and have a high specificity towards both NAD- and formate. FDHs from different organisms (bacteria, yeast, plants) function as dimers consisting of two identical subunits with a molecular weight from 35 to 50 kDa. The molecular mechanism of FDH is characterized by the direct transfer of a hydride ion from the substrate to the C4 atom of the nicotinamide ring of NAD- without additional proton transfer steps, which usually occurs in reactions catalyzed by related NAD--dependent dehydrogenases. Hence, the FDH-catalyzed reaction is a convenient model for studying the mechanism of hydride ion transfer in the active site of NAD--dependent hydrogenases by methods of quantum mechanics and molecular dynamics [3–5].

I G Shabalin

A.N. Bach Institute of Biochemistry RAS

K M Polyakov

A.N. Bach Institute of Biochemistry RAS; V.A. Engelhardt Institute of Molecular Biology RAS

V I Tishkov

A.N. Bach Institute of Biochemistry RAS

V O Popov

A.N. Bach Institute of Biochemistry RAS

Email: vpopov@inbi.ras.ru

  1. Tishkov V.I., Popov V.O. // Biochemistry Mosc. 2004. V. 69. № 11. P. 1252.
  2. Vinals C., Depiereux E., Feytmans E. // Biochem. Biophys. Res. Commun. 1993. V. 192. № 1. P. 182.
  3. Bandaria J.N., Dutta S., Hill S. E., Kohen A., Cheatum C.M. // J. Am. Chem. Soc. 2008. V. 130. № 1. P. 22.
  4. Castillo R., Oliva M., Marti S., Moliner V. // J. Phys. Chem. B. 2008. V. 112. № 32. P. 10012.
  5. Torres R.A., Schitt B., Bruice T.C. // J. Am. Chem. Soc. 1999. V. 121. № 36. P. 8164.
  6. Dauter Z., Lamzin V.S., Wilson K.S. // Curr. Opin. Struct. Biol. 1997. V. 7. № 5. P. 681.
  7. Rubach J.K., Plapp B.V. // Biochemistry. 2003. V. 42. № 10. P. 2907.
  8. Meijers R., Morris R.J., Adolph H.W., Merli A., Lamzin V.S., et al. // J. Biol. Chem. 2001. V. 276. № 12. P. 9316.
  9. Meijers R., Adolph H.W., Dauter Z., Wilson K. S., Lamzin V.S., et al. // Biochemistry. 2007. V. 46. № 18. P. 5446.
  10. Schlieben N.H., Niefind K., Muller J., Riebel B., Hummel W., et al. // J. Mol. Biol. 2005. V. 349. № 4. P. 801.
  11. Cameron A., Read J., Tranter R., Winter V. J., Sessions R.B., et al. // J. Biol. Chem. 2004. V. 279. № 30. P. 31429.
  12. Lamzin V.S., Dauter Z., Popov V.O., Harutyunyan E.H., Wilson K.S. // J. Mol. Biol. 1994. V. 236. № 3. P. 759.
  13. Schirwitz K., Schmidt A., Lamzin V.S. // Protein Sci. 2007. V.16. № 6. P. 1146.
  14. Shabalin I.G., Filippova E.V., Polyakov K.M., Sadykhov E.G., Safonova T.N., et al. // Acta Crystallogr. D Biol. Crystallogr. 2009. V. 65. № 12. P. 1315.
  15. Otwinowski Z., Minor W. // Methods in enzymology. 1997. V. 276. P. 307.
  16. Murshudov G.N., Vagin A.A., Dodson E.J. // Acta Crystallogr. D Biol. Crystallogr. 1997. V. 53. № 3. P. 240.
  17. Emsley P., Cowtan K. // Acta Crystallogr. D Biol. Crystallogr. 2004. V. 60. № 12. P. 2126.
  18. Laskowski R.A., MacArthur M.W., Moss D.S., Thornton J. M. // J. Appl. Cryst. 1993. V. 26. № 2. P. 283.
  19. Vaguine A.A., Richelle J., Wodak S.J. // Acta Crystallogr. D Biol. Crystallogr. 1999. V. 55. № 1. P. 191.
  20. Blanchard J.S., Cleland W.W. // Biochemistry. 1980. V. 19. № 15. P. 3543.
  21. Rotberg N.S., Cleland W.W. // Biochemistry. 1991. V. 30. № 16. P. 4068.
  22. Popov V.O., Lamzin V.S. // Biochem. J. 1994. V. 301. № 3. P. 625.
  23. Kahn K., Bruice T.C. // J. Am. Chem. Soc. 2001. V. 123. № 48. P. 11960.
  24. Engh R.A., Huber R. // Acta Crystallogr. A Found. Crystallogr. 1991. V. 47. № 4. P. 392.

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