Abstract
We expressed a putative β-galactosidase from Sulfolobus acidocaldarius in Escherichia coli and purified the recombinant enzyme using heat treatment and Hi-Trap ion-exchange chromatography. The resultant protein gave a single 57-kDa band by SDS-PAGE and had a specific activity of 58 U/mg. The native enzyme existed as a dimer with a molecular mass of 114 kDa by gel filtration. The maximum activity of this enzyme was observed at pH 5.5 and 90 ºC. The half-lives of the enzyme at 70, 80, and 90 ºC were 494, 60, and 0.2 h, respectively. The hydrolytic activity with p-nitrophenyl(pNP) substrates followed the order p-nitrophenyl-β-d-fucopyranoside > pNP-β-d-glucopyranoside > pNP-β-d-galactopyranoside > pNP-β-d-mannopyranoside > pNP-β-d-xylopyranoside, but not toward aryl-α-glycosides or pNP-β-l-arabinofuranoside. Thus, the enzyme was actually a β-glycosidase. The β-glycosidase exhibited transglycosylation activity with pNP-β-d-galactopyranoside, pNP-β-d-glucopyranoside, and pNP-β-d-fucopyranoside in decreasing order of activity, in the reverse order of its hydrolytic activity. The hydrolytic activity was higher toward cellobiose than toward lactose, but the transglycosylation activity was lower with cellobiose than with lactose.
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Sunna, A., Moracci, M., Rossi, M., & Antranikian, G. (1997). Extremophiles, 1, 2–13.
Bhatia, Y., Mishra, S., & Bisaria, V. S. (2002). Critical Reviews in Biotechnology, 22, 375–407.
Ducret, A., Trani, M., & Lortie, R. (2002). Biotechnology and Bioengineering, 77, 752–757.
Pisani, F. M., Rella, R., Raia, C. A., Rozzo, C., Nucci, R., Gambacorta, A., et al. (1990). European Journal of Biochemistry, 187, 321–328.
Atomi, H. (2005). Current Opinion in Chemical Biology, 9, 166–173.
Ciaramella, M., Pisani, F. M., & Rossi, M. (2002). Antonie Van Leeuwenhoek, 81, 85–97.
Dion, M., Fourage, L., Hallet, J. N., & Colas, B. (1999). Glycoconjugate Journal, 16, 27–37.
Kim, C. S., Ji, E. S., & Oh, D. K. (2004). Journal of Applied Microbiology, 97, 1006–1014.
Petzelbauer, I., Reiter, A., Splechtna, B., Kosma, P., & Nidetzky, B. (2000). European Journal of Biochemistry, 267, 5055–5066.
Ezaki, S., Miyaoku, K., Nishi, K., Tanaka, T., Fujiwara, S., Takagi, M., et al. (1999). Journal of Bioscience and Bioengineering, 88, 130–135.
Fourage, L., Dion, M., & Colas, B. (2000). Glycoconjugate Journal, 17, 377–383.
Grogan, D. W. (1991). Applied and Environmental Microbiology, 57, 1644–1649.
Ji, E. S., Park, N. H., & Oh, D. K. (2005). World Journal of Microbiology and Biotechnology, 21, 759–764.
Kim, C. S., Ji, E. S., & Oh, D. K. (2004). Biochemical and Biophysical Resarch Communications, 316, 738–743.
Park, H. Y., Kim, H. J., Lee, J. K., Kim, D., & Oh, D. K. (2008). World Journal of Microbiology and Biotechnology, 24, 1553–1558.
Coker, J. A., Sheridan, P. P., Loveland-Curtze, J., Gutshall, K. R., Auman, A. J., & Brenchley, J. E. (2003). Journal of Bacteriology, 185, 5473–5482.
Li, Y., Wang, H., Lu, L., Li, Z., Xu, X., & Xiao, M. (2008). Applied Biochemistry and Biotechnology, doi:10.1007/s12010-008-8310-4.
Hong, M. R., Kim, Y. S., Park, C. S., Lee, J. K., Kim, Y. S., & Oh, D. K. (2009). Journal of Bioscience and Bioengineering, 108, 36–40.
Kim, H. J., Park, A. R., Lee, J. K., & Oh, D. K. (2009). Biotechnology Letters, doi:10.1007/s10529-009-0018-1.
Petzelbauer, I., Zeleny, R., Reiter, A., Kulbe, K. D., & Nidetzky, B. (2000). Biotechnology and Bioengineering, 69, 140–149.
Kengen, S. W., Luesink, E. J., Stams, A. J., & Zehnder, A. J. (1993). European Journal of Biochemistry, 213, 305–312.
Henrissat, B. (1991). Biochemical Journal, 280(Pt 2), 309–316.
Bauer, M. W., & Kelly, R. M. (1998). Biochemistry, 37, 17170–17178.
Aguilar, C. F., Sanderson, I., Moracci, M., Ciaramella, M., Nucci, R., Rossi, M., et al. (1997). Journal of Molecular Biology, 271, 789–802.
Acknowledgments
This study was carried out with the support of ‘Forest Science and Technology Projects (Project No. S210707L010120)’ provided by Korea Forest Service and by the 21C Frontier Project for Microbial Genomics, Ministry of Education, Science, and Technology.
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Park, AR., Kim, HJ., Lee, JK. et al. Hydrolysis and Transglycosylation Activity of a Thermostable Recombinant β-Glycosidase from Sulfolobus acidocaldarius . Appl Biochem Biotechnol 160, 2236–2247 (2010). https://doi.org/10.1007/s12010-009-8705-x
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DOI: https://doi.org/10.1007/s12010-009-8705-x