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Contribution of conserved Glu255 and Cys289 residues to catalytic activity of recombinant aldehyde dehydrogenase from Bacillus licheniformis

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Abstract

Based on the sequence homology, we have modeled the three-dimensional structure of Bacillus licheniformis aldehyde dehydrogenase (BlALDH) and identified two different residues, Glu255 and Cys289, that might be responsible for the catalytic function of the enzyme. The role of these residues was further investigated by site-directed mutagenesis and biophysical analysis. The expressed parental and mutant proteins were purified by nickel-chelate chromatography, and their molecular masses were determined to be approximately 53 kDa by SDS-PAGE. As compared with the parental BlALDH, a dramatic decrease or even complete loss of the dehydrogenase activity was observed for the mutant enzymes. Structural analysis showed that the intrinsic fluorescence and circular dichroism spectra of the mutant proteins were similar to the parental enzyme, but most of the variants exhibited a different sensitivity towards thermal- and guanidine hydrochloride-induced denaturation. These observations indicate that residues Glu255 and Cys289 play an important role in the dehydrogenase activity of BlALDH, and the rigidity of the enzyme has been changed as a consequence of the mutations.

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Abbreviations

AEW:

average emission wavelength

ALDH:

aldehyde dehydrogenase

IPTG:

isopropyl β-D-thiogalactopy-ranoside

LB:

Luria-Bertani medium

References

  1. Perozich, J., Nicholas, H., Wang, B. C., Lindahl, R., and Hempel, J. (1999) Protein Sci., 8, 137–146.

    Article  PubMed  CAS  Google Scholar 

  2. Vasiliou, V., Pappa, A., and Petersen, D. (2000) Chem. Biol. Interact., 129, 1–19.

    Article  PubMed  CAS  Google Scholar 

  3. Sripo, T., Phongdara, A., Wanapu, C., and Caplan, A. B. (2002) Appl. Environ. Microbiol., 62, 171–179.

    Google Scholar 

  4. Chang, C., and Yoshida, A. (1994) Gene, 148, 331–336.

    Article  PubMed  CAS  Google Scholar 

  5. Fontaine, L., Meynial-Salles, I., Girbal, L., Yang, X., Croux, C., and Soucaille, P. (2002) J. Bacteriol., 184, 821–830.

    Article  PubMed  CAS  Google Scholar 

  6. Pappa, A., Estey, T., Manzer, R., Brown, D., and Vasiliou, V. (2003) Biochem. J., 376, 615–623.

    Article  PubMed  CAS  Google Scholar 

  7. Grochowski, L. L., Xu, H., and White, R. H. (2006) J. Bacteriol., 188, 2836–2844.

    Article  PubMed  CAS  Google Scholar 

  8. Bains, J., and Boulanger, M. J. (2008) J. Mol. Biol., 379, 597–608.

    Article  PubMed  CAS  Google Scholar 

  9. Cao, Y., Liao, L., Xu, X. W., Oren, A., and Wu, M. (2008) Extremophiles, 12, 849–854.

    Article  PubMed  CAS  Google Scholar 

  10. Jo, J. E., Mohan Raj, S., Rathnasingh, C., Selvakumar, E., Jung, W. C., and Park, S. (2008) Appl. Microbiol. Biotechnol., 81, 51–60.

    Article  PubMed  CAS  Google Scholar 

  11. Stiti, N., Adewale, I. O., Petersen, J., Bartels, D., and Kirch, H. H. (2011) Biochem. J., 434, 459–471.

    Article  PubMed  CAS  Google Scholar 

  12. Sophos, N. A., and Vasiliou, V. (2003) Chem. Biol. Interact., 143/144, 5–22.

    Article  Google Scholar 

  13. Vasiliou, V., and Nebert, D. W. (2005) Hum. Genom., 2, 138–143.

    CAS  Google Scholar 

  14. Kavanagh, K. L., Jornvall, H., Persson, B., and Oppermann, U. (2008) Cell. Mol. Life Sci., 65, 3895–3906.

    Article  PubMed  CAS  Google Scholar 

  15. Liu, Z. J., Sun, Y. J., Rose, J., Chung, Y. J., Hsiao, C. D., Chang, W. R., Kuo, I., Perozich, J., Lindahl, R., Hempel, J., and Wang, B. C. (1997) Nat. Struct. Biol., 4, 317–326.

    Article  PubMed  CAS  Google Scholar 

  16. Steinmetz, C. G., Xie, P., Weiner, H., and Hurley, T. D. (1997) Structure, 5, 701–711.

    Article  PubMed  CAS  Google Scholar 

  17. Johansson, K., el Almad, M., Ramaswamy, S., Hjelmqvist, L., Jornvall, H., and Eklund, H. (1998) Protein Sci., 7, 2106–2117.

    Article  PubMed  CAS  Google Scholar 

  18. Moore, S. A., Baker, H. M., Blythe, T. J., Kitson, K. E., Kitson, T. M., and Baker, N. (1998) Structure, 6, 1541–1551.

    Article  PubMed  CAS  Google Scholar 

  19. Lamb, A. L., and Newcomer, M. E. (1999) Biochemistry, 38, 6003–6011.

    Article  PubMed  CAS  Google Scholar 

  20. Cobessi, D., Tete-Favier, F., Marchal, S., Branlant, G., and Aubry, A. (1999) J. Mol. Biol., 290, 161–173.

    Article  PubMed  CAS  Google Scholar 

  21. Gonzalez-Segura, L., Rudino-Pinera, E., Munoz-Clares, R. A., and Horjales, E. (2009) J. Mol. Biol., 385, 542–557.

    Article  PubMed  CAS  Google Scholar 

  22. O’Donnell, A. G., Norris, J. R., Berkeley, R. C. W., Claus, D., Kaneko, T., Logan, N. A., and Nozaki, R. (1980) Int. J. Syst. Bacteriol., 30, 448–459.

    Article  Google Scholar 

  23. Xu, D., and Cote, J. C. (2003) Int. J. Syst. Evol. Microbiol., 53, 695–704.

    Article  PubMed  CAS  Google Scholar 

  24. Schallmey, M., Singh, A., and Ward, O. P. (2004) Can. J. Microbiol., 50, 1–17.

    Article  PubMed  CAS  Google Scholar 

  25. Ming, L. J., and Epperson, D. J. (2002) J. Inorg. Biochem., 91, 48–58.

    Article  Google Scholar 

  26. Murphy, T., Roy, I., Harrop, A., Dixon, K., and Keshavarz, T. (2007) J. Biotechnol., 131, 397–403.

    Article  PubMed  CAS  Google Scholar 

  27. Birrer, G. A., Cromwick, A. M., and Gross, R. A. (1994) Int. J. Biol. Macromol., 16, 265–275.

    Article  PubMed  CAS  Google Scholar 

  28. Zuo, R. (2007) Appl. Microbiol. Biotechnol., 76, 1245–1253.

    Article  PubMed  CAS  Google Scholar 

  29. Veith, B., Herzberg, C., Steckel, S., Feesche, J., Maurer, K. H., Ehrenreich, P., Baeumer, S., Henne, A., Liesegang, H., Merkl, R., Ehrenreich, A., and Gottschalk, G. (2004) J. Mol. Microbiol. Biotechnol., 7, 204–211.

    Article  PubMed  CAS  Google Scholar 

  30. Lo, H. F., and Chen, Y. J. (2010) Mol. Biotechnol., 46, 157–167.

    Article  PubMed  CAS  Google Scholar 

  31. Nimlos, M. R., Matthews, J. F., Crowley, M. F., Walker, R. C., Chukkapalli, G., Brady, J. W., Adney, W. S., Cleary, J. M., Zhong, L., and Himmel, M. E. (2007) Protein Eng. Des. Sel., 20, 179–187.

    Article  PubMed  CAS  Google Scholar 

  32. Laemmli, U. K. (1970) Nature, 227, 680–685.

    Article  PubMed  CAS  Google Scholar 

  33. Bradford, M. M. (1976) Anal. Biochem., 72, 248–254.

    Article  PubMed  CAS  Google Scholar 

  34. Leal, N. A., Havemann, G. D., and Bobik, T. A. (2003) Arch. Microbiol., 180, 353–361.

    Article  PubMed  CAS  Google Scholar 

  35. Royer, C. A., Mann, C. J., and Mathews, C. R. (1995) Protein Sci., 2, 1844–1852.

    Article  Google Scholar 

  36. Pace, C. N. (1990) Trends Biotechnol., 8, 93–98.

    Article  PubMed  CAS  Google Scholar 

  37. Cobessi, D., Tete-Favier, F., Marchal, S., Branlant, G., and Aubry, A. (2000) J. Mol. Biol., 300, 141–152.

    Article  PubMed  CAS  Google Scholar 

  38. Marchal, S., Rahuel-Clermont, S., and Branlant, G. (2000) Biochemistry, 39, 3327–3335.

    Article  PubMed  CAS  Google Scholar 

  39. Li, N., Ma, D. L., Liu, X., Wu, L., Chu, X., Wong, K. Y., and Li, D. (2007) Protein J., 26, 569–576.

    Article  PubMed  CAS  Google Scholar 

  40. Zhao, L., Liu, D., Zhang, Q., Zhang, S., Wan, J., and Xiao, W. (2008) FEMS Microbiol. Lett., 277, 37–43.

    Article  Google Scholar 

  41. Lo, H. F., Chen, H. L., Yen, S. Y., Ong, P. L., Chang, W. S., and Chang, C. T. (2010) Biologia, 65, 399–407.

    Article  CAS  Google Scholar 

  42. DeLano, W. L. (2002) The PyMOL Molecular Graphics System, DeLano Scientific, Palo Alto, CA, USA, http://www.pymol.org/

    Google Scholar 

  43. Thornton, J. M., Orengo, C. A., Todd, A. E., and Pearl, F. M. (1999) J. Mol. Biol., 293, 333–342.

    Article  PubMed  CAS  Google Scholar 

  44. Feldman, R. I., and Weiner, H. (1972) J. Biol. Chem., 247, 267–272.

    PubMed  CAS  Google Scholar 

  45. Wang, X., and Weiner, H. (1995) Biochemistry, 34, 237–243.

    Article  PubMed  CAS  Google Scholar 

  46. Vedadi, M., Szittner, R., Smillie, L., and Meighen, E. (1995) Biochemistry, 34, 16725–16732.

    Article  PubMed  CAS  Google Scholar 

  47. Ahvazi, B., Coulombe, R., Delarge M., Vedadi, M., Zhang, L., Meihgen, E., and Vrielink, A. (2000) Biochem. J., 349, 853–861.

    PubMed  CAS  Google Scholar 

  48. Gruez, A., Roig-Zamboni, V., Grisel, S., Salomoni, A., Valencia, C., Campanacci, V., Tegoni, M., and Cambillau, C. (2004) J. Mol. Biol., 343, 29–41.

    Article  PubMed  CAS  Google Scholar 

  49. Inagaki, E., Ohshima, H., Takahashi, C., Yokoyama, S., and Tahirov, T. H. (2006) J. Mol. Biol., 362, 490–501.

    Article  PubMed  CAS  Google Scholar 

  50. Tsybovsky, Y., Donato, H., Krupenko, N. I., Davies, C., and Krupenko, S. A. (2007) Biochemistry, 46, 2917–2929.

    Article  PubMed  CAS  Google Scholar 

  51. Tylichova, M., Kopecny, D., Morera, S., Briozzo, P., Lenobel, R., Snegaroff, J., and Sebela, M. (2010) J. Mol. Biol., 396, 870–882.

    Article  PubMed  CAS  Google Scholar 

  52. Di Constanzo, L., Gomez, G. A., and Christiaanson, D. W. (2007) J. Mol. Biol., 366, 481–493.

    Article  Google Scholar 

  53. Perez-Miller, S. J., and Hurley, T. D. (2003) Biochemistry, 42, 7100–7109.

    Article  PubMed  CAS  Google Scholar 

  54. Larson, H. N., Zhou, J., Chen, Z., Stamler, J. S., Weiner, H., and Hurley, T. D. (2007) J. Biol. Chem., 282, 12940–12950.

    Article  PubMed  CAS  Google Scholar 

  55. Pohl, E., Brunner, N., Wilmanns, M., and Hensel, R. (2002) J. Biol. Chem., 277, 19938–19945.

    Article  PubMed  CAS  Google Scholar 

  56. Lorentzen, E., Hensel, R., Knura, T., Ahmed, H., and Pohl, E. (2004) J. Mol. Biol., 341, 815–828.

    Article  PubMed  CAS  Google Scholar 

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Authors and Affiliations

  1. Department of Bioagricultural Science, National Chiayi University, 300 Syuefu Road, Chiayi City, 60004, Taiwan

    Yen-Chung Lee & Den-Tai Lin

  2. Department of Biochemical Science and Technology, National Chiayi University, 300 Syuefu Road, Chiayi City, 60004, Taiwan

    Ping-Ling Ong

  3. Department of Food Science and Technology, Hungkuang University, Shalu, Taichung City, Taiwan

    Hsiang-Ling Chen & Huei-Fen Lo

  4. Department of Applied Chemistry, National Chiayi University, 300 Syuefu Road, Chiayi City, 60004, Taiwan

    Long-Liu Lin

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  1. Yen-Chung Lee
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Correspondence to Huei-Fen Lo or Long-Liu Lin.

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Published in Biokhimiya, 2011, Vol. 76, No. 11, pp. 1532–1541.

Originally published in Biochemistry (Moscow) On-Line Papers in Press, as Manuscript BM11-171, September 11, 2011.

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Lee, YC., Lin, DT., Ong, PL. et al. Contribution of conserved Glu255 and Cys289 residues to catalytic activity of recombinant aldehyde dehydrogenase from Bacillus licheniformis . Biochemistry Moscow 76, 1233–1241 (2011). https://doi.org/10.1134/S0006297911110058

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  • DOI: https://doi.org/10.1134/S0006297911110058

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