Advertisement

Applied Biochemistry and Biotechnology

, Volume 170, Issue 3, pp 639–653 | Cite as

Recombinant S-Adenosylhomocysteine Hydrolase from Thermotoga maritima: Cloning, Overexpression, Characterization, and Thermal Purification Studies

  • J. D. Lozada-RamírezEmail author
  • A. Sánchez-Ferrer
  • F. García-Carmona
Article

Abstract

S-Adenosylhomocysteine hydrolase (SAHase) encoded by sahase gene is a determinant when catalyzing the reversible conversion of adenosine and homocysteine to S-adenosylhomocysteine in most living organisms. The sahase gene was isolated from the genome of the highly thermostable anaerobic bacteria Thermotoga maritima, and then it was cloned, characterized, overexpressed using Escherichia coli, and partially purified by thermal precipitation. The thermal purification of the recombinant SAHase resulted in changes in the circular dichroism spectra. As a result of this analysis, it was possible to determine the structural changes in the composition of the α-helix and β-sheet content of the recombinant enzyme after purification. Moreover, a predicted secondary structure and 3D structural model was rendered by comparative molecular modeling to further understand the molecular function of this protein including its attractive biotechnological use.

Keywords

S-Adenosylhomocysteine hydrolase S-Adenosyl-l-homocysteine Thermal purification Thermotoga maritima Structural modeling 

Abbreviations

SAHase

S-Adenosyl-l-homocysteine hydrolase

rSAHase

Recombinant S-adenosyl-l-homocysteine hydrolase

SAH

S-Adenosyl-l-homocysteine

ADO

Adenosine

Hcy

Homocysteine

Cys

Cysteine

CD

Circular dichroism

SAM

S-Adenosylmethionine

sahase

S-Adenosylhomocysteine hydrolase coding gene

Notes

Acknowledgments

This study was partially supported by Spanish grants from MINECO-FEDER (BIO2010-22225-C02-01) and from Programa de Ayuda a Grupos de Excelencia de la Región de Murcia, Fundación Séneca (04541/GERM/06, Plan Regional de Ciencia y Tecnología 2007–2010), and by Proyecto CONACYT Ciencia Básica 2009–2012 (CB-133949).

Supplementary material

12010_2013_218_MOESM1_ESM.jpg (104 kb)
Additional material 1 (A) 3D structural model of a single unit of SAHase from Thermotoga maritima. Alpha-helix are indicated in black, beta-strand in gray, and random coils in white. (B) Amino acid residues of the conserved domains implied in SAHase catalytic activity (Ado binding), and in NAD+ binding. Amino acid residues are discussed in the text. (JPEG 103 kb)
12010_2013_218_MOESM2_ESM.jpg (175 kb)
Additional material 2 Ribbon diagrams of the tetrameric structure of rat SAHase (A) and the theoretical structure of T. maritima SAHase. (B) Enlargements. (C) and (D) Amino acid residues which are involved in the network interactions in the central channel of rat SAHase and T. maritima SAHase, respectively. Amino acid residues are discussed in the text. (JPEG 175 kb)

References

  1. 1.
    de la Haba, G., & Cantoni, G. L. (1959). J Parasit, 234, 603–608.Google Scholar
  2. 2.
    Ueland, P. M. (1982). Pharmacological Reviews, 34, 223–253.Google Scholar
  3. 3.
    Chiang, P. K., Gordon, R. K., Tal, J., Zeng, G. C., Doctor, B. P., Pardhasaradhi, K., et al. (1996). The FASEB Journal, 10, 471–480.Google Scholar
  4. 4.
    Yin, D., Yang, X., Hu, Y., Kuczera, K., Schowen, R. L., Borchardt, R. T., et al. (2000). Biochemistry, 39, 9811–9818.CrossRefGoogle Scholar
  5. 5.
    Walker, R. D., & Duerre, J. A. (1975). Canadian Journal of Biochemistry, 53, 312–319.CrossRefGoogle Scholar
  6. 6.
    Wnuk, S. F. (2001). Mini Rev. Medicinal Chemistry, 1, 307–316.Google Scholar
  7. 7.
    Kajander, E. O., & Raina, A. M. (1981). Biochemical Journal, 193, 503–512.Google Scholar
  8. 8.
    Henderson, D. M., Hanson, S., Allen, T., Wilson, K., Coulter-Karis, D. E., Greenberg, M. L., et al. (1992). Molecular and Biochemical Parasitology, 53, 169–183.CrossRefGoogle Scholar
  9. 9.
    Creedon, K. A., Rathod, P. K., & Wellems, T. E. (1994). Journal of Biological Chemistry, 269, 16364–16370.Google Scholar
  10. 10.
    Cantoni, G. L. (1986). Biological methylation and drug design. In R. T. Borchardt, C. R. Creveiling, & P. M. Ueland (Eds.), The centrality of S-adenosylhomocysteinase in the regulation of the biological utilization of S-adenosylmethionine (pp. 227–238). Totowa: Humana.Google Scholar
  11. 11.
    Hayden, D. M., Rolletschek, H., Borisjuk, L., Corwin, J., Kliebenstein, D. J., Grimberg, A., et al. (2011). The Plant Journal, 67, 1018–1028.CrossRefGoogle Scholar
  12. 12.
    Siu, K. K., Asmus, K., Zhang, A. N., Horvatin, C., Li, S., Liu, T., et al. (2011). Journal of Structural Biology, 173, 86–98.CrossRefGoogle Scholar
  13. 13.
    Choi, J., Choi, D., Lee, S., Ryu, C. M., & Hwang, I. (2011). Trends in Plant Science, 16, 388–394.CrossRefGoogle Scholar
  14. 14.
    Keller, W., & Bekkaoui, F. (2009). Botany, 87, 519–525.CrossRefGoogle Scholar
  15. 15.
    Wu, X., Li, F., Kolenovsky, A., Caplan, A., Cui, Y., Cutler, A., et al. (2009). Botany, 87, 571–584.CrossRefGoogle Scholar
  16. 16.
    Masuta, C., Tanaka, H., Uehara, K., Kuwata, S., Koiwai, A., & Noma, M. (1995). Proceedings of the National Academy of Sciences of the United States of America, 92, 6117–6121.CrossRefGoogle Scholar
  17. 17.
    Hendricks, C. L., Ross, J. R., Pichersky, E., Noel, J. P., & Zhou, Z. S. (2004). Analytical Biochemistry, 326, 100–105.CrossRefGoogle Scholar
  18. 18.
    Edwards, A. L., Reyes, F. E., Héroux, A., & Batey, R. T. (2010). RNA, 16, 2144–2155.CrossRefGoogle Scholar
  19. 19.
    Collazo, E., Couture, J. F., Bulfer, S., & Trievel, R. C. (2005). Analytical Biochemistry, 342, 86–92.CrossRefGoogle Scholar
  20. 20.
    Palmer, N. A., Sattler, S. E., Saathoff, A. J., & Sarath, G. (2010). Journal of Agricultural and Food Chemistry, 12, 5220–5226.CrossRefGoogle Scholar
  21. 21.
    Bujnicki, J. M., Prigge, S. T., Cardinha, D., & Chiang, P. K. (2003). Proteins, 52, 624–632.CrossRefGoogle Scholar
  22. 22.
    Kim, B. G., Chun, T. G., Lee, H. Y., & Snapper, M. L. (2009). Bioorganic & Medicinal Chemistry, 15, 6707–6714.CrossRefGoogle Scholar
  23. 23.
    Carlucci, F., Tabucchi, A., Aiuti, A., Rosi, F., Floccari, F., Pagani, R., et al. (2003). Clinical Chemistry, 49, 1830–1838.CrossRefGoogle Scholar
  24. 24.
    Baric, I., Fumic, K., Glenn, B., Cuk, M., Schulze, A., Finkelstein, J. D., et al. (2004). Proceedings of the National Academy of Sciences of the United States of America, 23, 4234–4239.CrossRefGoogle Scholar
  25. 25.
    Liszka, M. J., Clark, M. E., Schneider, E., & Clark, D. S. (2012). Annual Review of Chemical and Biomolecular Engineering, 3, 77–102.CrossRefGoogle Scholar
  26. 26.
    Porcelli, M., Fusco, S., Inizio, T., Zappia, V., & Cacciapuoti, G. (2000). Protein Expression and Purification, 18, 27–35.CrossRefGoogle Scholar
  27. 27.
    Porcelli, M., Moretti, M. A., Concilio, L., Forte, S., Merlino, A., Graziano, G., et al. (2005). Proteins, 58, 815–825.CrossRefGoogle Scholar
  28. 28.
    Marino, G., Nitti, G., Arnone, M. I., Sannia, G., Gambacorta, A., & De Rosa, M. (1998). Journal of Biological Chemistry, 263, 12305–12309.Google Scholar
  29. 29.
    Jones, C. E., Fleming, T. M., Cowan, D. A., Littlechild, J. A., & Piper, P. W. (1995). European Journal of Biochemistry, 233, 800–808.CrossRefGoogle Scholar
  30. 30.
    Kumar, S., & Nussinov, R. (2001). Cellular and Molecular Life Sciences, 58, 1216–1233.CrossRefGoogle Scholar
  31. 31.
    Fabry, S., & Hensel, R. (1987). European Journal of Biochemistry, 165, 147–155.CrossRefGoogle Scholar
  32. 32.
    Simpson, H. D., Haufler, U. R., & Daniel, R. M. (1991). Biochemical Journal, 277, 413–417.Google Scholar
  33. 33.
    Koch, R., Canganella, F., Hippe, H., Jahnke, K. D., & Antranikian, G. (1997). Applied and Environmental Microbiology, 63, 1088–1094.Google Scholar
  34. 34.
    Klingeberg, M., Galunsky, B., Sjoholm, C., Kasche, V., & Antranikian, G. (1995). Applied and Environmental Microbiology, 61, 3094–3104.Google Scholar
  35. 35.
    Wassenberg, D., Liebl, W., & Jaenicke, R. (2000). Journal of Molecular Biology, 295, 279–288.CrossRefGoogle Scholar
  36. 36.
    Andreotti, G., Cubellis, M. V., Nitti, G., Sannia, G., Mai, X., Adams, M. W. W., et al. (1995). Biochimica et Biophysica Acta, 1247, 90–96.CrossRefGoogle Scholar
  37. 37.
    Vieille, C., & Zeikus, G. J. (2001). Microbiology and Molecular Biology Reviews, 65, 1–43.CrossRefGoogle Scholar
  38. 38.
    Sreerama, N., Venyaminov, S. Y., & Woody, R. W. (2000). Analytical Biochemistry, 287, 243–251.CrossRefGoogle Scholar
  39. 39.
    Bradford, M. M. (1976). Analytical Biochemistry, 72, 248–254.CrossRefGoogle Scholar
  40. 40.
    Andrade, M. A., Chacón, P., Merelo, J. J., & Morán, F. (1993). Protein Engineering, 6, 383–390.CrossRefGoogle Scholar
  41. 41.
    Merelo, J. J., Andrade, M. A., Prieto, A., & Morán, F. (1994). Neurocomputing, 6, 443–454.CrossRefGoogle Scholar
  42. 42.
    Deleage, G., & Roux, B. (1987). Protein Engineering, 1, 289–294.CrossRefGoogle Scholar
  43. 43.
    Geourjon, C., & Deleage, G. (1994). Protein Engineering, 7, 157–164.CrossRefGoogle Scholar
  44. 44.
    Guermeur, Y., Geourjon, C., Gallinari, P., & Deleage, G. (1999). Bioinformatics, 15, 413–421.CrossRefGoogle Scholar
  45. 45.
    King, R. D., & Sternberg, M. J. (1996). Protein Science, 5, 2298–2310.CrossRefGoogle Scholar
  46. 46.
    Peitsch, M. C., Wells, T. N., Stampf, D. R., & Sussman, J. L. (1995). Trends in Biochemical Sciences, 20, 82–84.CrossRefGoogle Scholar
  47. 47.
    Guex, N., & Peitsch, M. C. (1997). Electrophoresis, 18, 2714–2723.CrossRefGoogle Scholar
  48. 48.
    Schwede, T., Kopp, J., Guex, N., & Peitsch, M. C. (2003). Nucleic Acids Research, 31, 3381–3385.CrossRefGoogle Scholar
  49. 49.
    Lozada-Ramírez, J. D., Martínez-Martínez, I., Sánchez-Ferrer, A., & García-Carmona, F. (2006). Journal of Biochemical and Biophysical Methods, 67, 131–140.CrossRefGoogle Scholar
  50. 50.
    Porcelli, M., Cacciapuoti, G., Fusco, S., Iacomino, G., Gambacorta, A., De Rosa, M., et al. (1993). Biochimica et Biophysica Acta, 1164, 179–188.CrossRefGoogle Scholar
  51. 51.
    Huber, R., Langworthy, T. A., Köning, H., Thomm, M., Woese, C. R., Sleytr, U. B., et al. (1986). Archives of Microbiology, 144, 324–333.CrossRefGoogle Scholar
  52. 52.
    Bouthier de la Tour, C., Portemer, C., Kaltoum, H., & Duguet, M. (1998). Journal of Bacteriology, 180, 274–281.Google Scholar
  53. 53.
    Yamada, T., Takata, Y., Komoto, J., Gomi, T., Ogawa, H., Fujioka, M., et al. (2005). International Journal of Biochemistry and Cell Biology, 37, 2417–2435.CrossRefGoogle Scholar
  54. 54.
    Tanaka, N., Nakanishi, M., Kusakabe, Y., Shiraiwa, K., Yabe, S., Ito, Y., et al. (2004). Journal of Molecular Biology, 343, 1007–1017.CrossRefGoogle Scholar
  55. 55.
    Bethin, K. E., Petrovic, N., & Ettinger, M. J. (1995). Journal of Biological Chemistry, 270, 20698.CrossRefGoogle Scholar
  56. 56.
    Aguilar, C. F., Sanderson, I., Moracci, M., Ciaramella, M., Nucci, R., Rossi, M., et al. (1997). Journal of Molecular Biology, 271, 789–802.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • J. D. Lozada-Ramírez
    • 1
    Email author
  • A. Sánchez-Ferrer
    • 2
  • F. García-Carmona
    • 2
  1. 1.Department of Chemical and Biological Sciences, School of SciencesUniversidad de las Américas PueblaSanta Catarina Mártir CholulaMéxico
  2. 2.Department of Biochemistry and Molecular Biology-A, Faculty of BiologyUniversity of MurciaMurciaSpain

Personalised recommendations