Food Science of Animal Resources (한국축산식품학회지)

Korean Society for Food Science of Animal Resources (한국축산식품학회)
권호 표지
DOI QR코드

DOI QR Code

Purification and Characterization of α-Galactosidase from Lactobacillus salivarius subsp. salivarius Nam27

  • Bae, Hyoung-Cchurl (Division of Animal Science & Resources, Chungnam National University) ;
  • Choi, Jong-Woo (Department of Food Science, University of Wisconsin) ;
  • Nam, Myoung-Soo (Division of Animal Science & Resources, Chungnam National University)
  • Published : 2007.03.31
Download PDF
⟨ Previous Next ⟩

Abstract

Lactobacillus salivarius subsp. salivarius CNU27 possessed a high level of ${\alpha}$-galactosidase activity. Purified ${\alpha}$-galactosidase was obtained after sonication of harvested cell pellet followed by DEAE-Sephadex A-50 and Mono Q anion exchange chromatography. The specific activity of the purified enzyme was 8,994 units/mg protein which is 17.09 times higher than that in crude extract. The native enzyme was a monomer with a molecular mass of 56,397.1 dalton. The optimum temperature and pH for the enzyme were $40^{\circ}C$ and 6.0, respectively. The enzyme was stable between 25 and $50^{\circ}C$. However, ${\alpha}$-galactosidase activity was lost rapidly below pH 4.5 and above pH 8.5. The enzyme activity decreased to 6.73% and 4.30% of the original activity by addition of $Cu^{2+}$ and $Hg^{2+}$, respectively. Other metal compounds did not affect the enzyme activity significantly. The enzyme liberated galactose from melibiose, raffinose, and stachyose. The rate of substrates hydrolysis was measured by HPLC. Raffinose, stachyose and melibiose were completely decomposed after 24 hr at $40^{\circ}C$.

Keywords

References

  1. Aduse-Opoku, J., Tao, L., Ferretti, J. J., and Russell, R. R. B. (1991) Biochemical and genetic analysis of Streptococcus mutans a-galactosidase. J. Gen. Microbiol. 137,2271-2272 https://doi.org/10.1099/00221287-137-9-2271
  2. Bae, B. C., Nam, M. S., and Lee, J. Y. (2002) Probiotic characterization of acid- and bile-tolerant lactobacillus salivarius subsp. salivarius from korean faeces. Asian-Aust. J. Anim. Sci. 15, 1798-1807 https://doi.org/10.5713/ajas.2002.1798
  3. Bae, B. C., Choi, S. B., and Nam, M. S. (2001) Isolation and identification of acid- and bile-tole rant Lactobacillus salivarius subsp. salivarius from human faeces. Asian-Aust. J. Anim. Sci. 14, 1170-1178 https://doi.org/10.5713/ajas.2001.1170
  4. Bradford, M. M. (1976) A rapid and sensitive method for quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72, 248-254 https://doi.org/10.1016/0003-2697(76)90527-3
  5. Civas, A., Eberhard, R., Dizet, P. L., and Petek, F. (1984) Glycosidases induced in Aspergillus tamarii mycelial $\alpha$-Dgalactosidases. J. Biochem. 219, 849-855 https://doi.org/10.1042/bj2190849
  6. Gao, Z., and Schaffer, A. A. (1999) A novel alkaline $\alpha$-galactosidase from melon fruit with a substrate preference for raffinose. Plant Physiol. 119,979-987 https://doi.org/10.1104/pp.119.3.979
  7. Garro, M. S., de Giori, O. S., de Valdez, O. F., and Oliver, G. (1993) Characterization of alpha-galactosidase from Lactobacillus fermentum. J. Appl. Bacteriol. 75, 485-488 https://doi.org/10.1111/j.1365-2672.1993.tb02805.x
  8. Gherardini, F., Babcock, M., and Salyers, A. A. (1985) Purification and characterization of two $\alpha$-galactosidases associated with catabolism of guar gum and other $\alpha$-galactosides by Bacteroides ovatus. J. Bacteriol, 161, 500-506
  9. Jin, F., Li, Y., Zhang, C., and Yu, H. (2001) Thermostable aamylase and $\alpha$-galactosidase production from the thermophilic and aerobic Bacillus sp. JF strain. Process Biochem. 36, 559-564 https://doi.org/10.1016/S0032-9592(00)00247-8
  10. Keller, F., and Pharr, D. M. (1996) Metabolism of carbohydrates in sink and sources: galactosyl-sucrose oligosaccharides. In: Photoassimilate Distribution in Plants and Crops. Marcel Dekker, New York. pp. 157-183
  11. Kotwal, S. M., Gote, M. M., Khan, M. I., and Khire, J. M. (1999) Production, purification and characterization of a constitutive intracellular $\alpha$-galactosidase from the thermophilic fungus Humicola sp. J. Ind. Microbiol. & Biotechnol. 23, 661-667 https://doi.org/10.1038/sj.jim.2900680
  12. Leder, S., Hartmeier, W., and Marx, S. P. (1999) Alpha-galactosidase of Bifidobacterium adolescentis DSM 20083. Curr. Microbiol. 38, 101-106 https://doi.org/10.1007/s002849900411
  13. Li, Y. T., Li, S. C. C., and Shetlar, M. R. (1963) $\alpha$-Galactosidase from Diplococcus pneumoniae. Arch. Biochem. Biophys. 103,436-442 https://doi.org/10.1016/0003-9861(63)90434-X
  14. Mitsutomi, M., and Ohtakara, A. (1988) Isolation and identification of oligosaccharides produced from raffinose by transgalactosylation reaction of thermostable $\alpha$-galactosidase from Pycnoporus cinnabarinus. Agric. Biol. Chem. 52, 2305-2311 https://doi.org/10.1271/bbb1961.52.2305
  15. Murphy, R. A., and Power, R. F. O. (2002) Expression of an $\alpha$-galactosidase from Saccharomyces cerevisiae in Aspergillus awamori and Aspergillus oryzae. J. Industrial Microbiol. & Biotechnol. 28,97-102 https://doi.org/10.1038/sj/jim/7000217
  16. Nagao, Y., Nakada, T., Imoto, T., Shimamoto, M., Sakai, T., Tsuda, S., and Tsuchiya, T. (1988) Purification and analysis of the structure of $\alpha$-galactosidase from Escherichia coli. Biochem. Biophys. Res. Commun. 151,236-241 https://doi.org/10.1016/0006-291X(88)90584-0
  17. Ohshima, T., Murray, O. J., Swaim, W. D., Longenecker, G., Quirk, J. M., Cardarelli, C. O., Sugimoto, Y., Pastan, I., Gottesman, M. M., Brady, R. O., and Kulkarni, A. B. (1997) $\alpha$-Galactosidase A deficient mice: a model of Fabry disease. In: Proceedings of the National Academy of Sciences of the United States of America, pp. 2540-2544
  18. Roy, D., Blanchette, L., Savoie, L., Ward, P., and Chevalier, P. (1992) Alpha- and beta-galactosidase properties of Bifidobacterium infantis. Milchwissenschaft. 47, 18-21
  19. Savel'ev, A. N., Ibatyllin, F. M., Eneyskaya, E. V., Kachurin, A. M., and Neustroev, K. N. (1996). Enzymatic properties of $\alpha$-galactosidase from Trichoderma reesei. Carbohydrate Research, 296,261-273 https://doi.org/10.1016/S0008-6215(96)00248-0
  20. Shabalin, K. A., Kulminskaya, A. A., Savel'ev, A. N., ShishIyannikov, S. M., and Neustroev, K. N. (2002) Enzymatic properties of $\alpha$-galactosidase from Trichoderma reesei in the hydrolysis of galactooligosaccharides. Enzyme Microbial Technol. 30, 231-239 https://doi.org/10.1016/S0141-0229(01)00482-3
  21. Silvestroni, A., Connes, C., Sesma, F., De-Giori, O. S., and Piard, J. C. (2002) Characterization of the melA Locus for alpha-Galactosidase in Lactobacillus plantarum. Appl. Environ. Microbiol. 68,5464-5471 https://doi.org/10.1128/AEM.68.11.5464-5471.2002
  22. Tzortzis, G., Jay, A. J., Baillon, M. L. A., Gibson, O. R., and Rastall, R. A. (2003) Synthesis of $\alpha$-galacto-oligosaccharides with $\alpha$-galactosidase from Lactobacillus reuteri of canine origin. Appl. Microbiol. Biotechnol. 63, 286-292 https://doi.org/10.1007/s00253-003-1426-0
  23. Yeo, N. I., Lee, S. K., and Ji, O. E. (1993) Characterization of $\alpha$-galactosidase from Bifidobacterium sp. Int-57. Kor. J. Food Sci. Technol. 25, 689-693
  24. Zeilinger, S., Kristufek, D., Inci, A. A., Hodits, R., and Kubicek, C. P. (1993) Conditions of formation, purification and characterization of an $\alpha$-galactosidase of Trichoderma reesei RUT-30. Appl. Environ. Microbiol. 59, 1347-1353