Skip to main content

Advertisement

Dealing with different methods for Kluyveromyces lactis β-galactosidase purification

Article metrics

Abstract

Several micro-scale chromatography-based procedures for purification of the β-galactosidase from the yeast Kluyveromyces lactis were assayed. Purified enzyme was suitable to be used as antigen to induce polyclonal antibodies production. Specific staining of non-denaturing PAGE gels with chromogenic substrates allowed the determination of the number of subunits forming the native enzyme.

References

  1. 1.

    González Siso, M.I. 1996. The biotechnological utilization of cheese whey: a review. Biores.Technol. 57, 1–11.

  2. 2.

    González Siso, M.I. 1994. β-galactosidase production by Kluyveromyces lactis on milk whey: batch versus fed-batch cultures. Proc. Biochem. 29, 565–568

  3. 3.

    Becerra, M. and González Siso, M.I. 1996. Yeast β-galactosidase in solid-state fermentations. Enz. Microb.Technol. 19, 33–44.

  4. 4.

    González Siso, M.I., Freire Picos, A., Ramil, E., RodrÍguez Belmonte, E., RodrÍguez Torres, A. and Cerdán, E. 1994. Covalent immobilization of β-galactosidase on corn grits. A system for lactose hydrolysis without diffusional resistance. Proc. Biochem. 29, 7–12

  5. 5.

    González Siso, M.I., Cerdán, E., Freire Picos, M. A., RodrÍguez Belmonte, E. and RodrÍguez Torres, A.M. 1992. Permeabilization of Kluyveromyces lactis cells for milk whey saccharification: a comparison of different treatments. Biotechnol.Techniques 6, 289–292.

  6. 6.

    González Siso, M.I. and Suárez Doval, S. 1994. Kluyveromyces lactis immobilization on corn grits for milk whey lactose hydrolysis. Enzyme Microb. Technol. 16, 303–310.

  7. 7.

    Becerra, M., Cerdán, E. and González Siso, M.I. 1997. Heterologous Kluyveromyces lactis β-galactosidase production and release by Saccharomyces cerevisiae osmotic-remedial thermosensitive autolytic mutants. Biochim. Biophys. Acta. 1335, 235–241.

  8. 8.

    Becerra, M., Cerdán, E. and González Siso, M.I. 1998. Micro-scale purification of β-galactosidase from Kluyveromyces lactis reveals that dimeric and tetrameric forms are active. Biotechnol.Techniques 12, 253–256.

  9. 9.

    Nader de Macias, M.E., Perdigon, G., Oliver, G. and A. A. P. de Ruiz Holgado. 1985. Immunological relationships among β-galactosidases in members of the genus Lactobacillus. Int. J. Sys. Bacteriol. 35, 103–110.

  10. 10.

    Guarante, L. 1983. Yeast promoters and LacZ fusions designed to study expression of cloned genes in yeasts. Meth. Enzymol. 101, 181–189.

  11. 11.

    Inchaurrondo, V. A., Yantorno, O.M. and Voget, C.E. 1994. Yeast growh and β-galactosidase production during aerobic batch cultures in lactose limited synthetic medium. Proc. Biochem. 29, 47–54.

  12. 12.

    Bradford, M.M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72, 248–254.

  13. 13.

    Laemmli, U. K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680–685.

  14. 14.

    Neuhoff, V., Stamm, R. and Eibk, M. 1985. Clear background and highly sensitive protein staining with Coomassie blue dyes in polyacrylamide gels: a systematic analysis. Electrophoresis 6, 427–448.

  15. 15.

    Dickson, R.C., Dickson, L.R. and Markin, J.S. 1979. Purification and properties of an inducible β-galactosidase isolated from the yeast Kluyveromyces lactis. J. Bacteriol. 137, 51–61.

  16. 16.

    Erickson, R.P. and Steers, J.R. E. 1970. Comparative study of isoenzyme formation of bacterial β-Galactosidase. J. Bacteriol. 102, 79–84.

  17. 17.

    Brady, D., Marchant, R., McHale, L. and McHale, A.P. 1995. Isolation and partial characterization of β-galactosidase activity produced by a thermotolerant strain of Kluyveromyces marxianus during growth on lactose-containing media. Enzyme Microb. Technol. 17, 696–699.

  18. 18.

    Pritchard, D.I., Maizels, R.M., Behnke, J.M. and Appleby, P. 1984. Stage-specific antigens of Nematospiroides dubius. Immunology 53, 325–335.

  19. 19.

    Iglesias, R., Leiro, J., Ubeira, F.M., Santamarina, M.T. and Sanmartín, M.L. 1993. Anisakis simplex: antigen recognition and antibody production in experimentally infected mice. Parasite Immunol. 15, 243–250.

Download references

Author information

Correspondence to M. I. González Siso.

Rights and permissions

Reprints and Permissions

About this article

Keywords

  • Lactase
  • Cheese Whey
  • Purification Factor
  • Biological Procedure
  • Crude Protein Extract