α-GALACTOSIDASE OF ASPERGILLUS NIGER: PURIFICATION AND PROPERTIES

N. V. Borzova, L. D. Varbanets


DOI: http://dx.doi.org/10.30970/sbi.0101.003

Abstract


Highly purified α-galactosidase (specific activiys 25 U/mg protein) has been isolated from the cultural liquid of Aspergillus niger. The enzyme is thermo- and pH-stable, with pH-optimum 4.1, and temperature optimum 60 °C. Km and Vmax for nitrophenyl substrate was shown to be 1.19 mM and 25 µmol/min/mg protein, respectively. α-Galactosidase was inhibited by the product of reaction D-galactose (Ki – 6.2 × 10-2M). The enzyme displayed narrow specificity towards glycon. It proved to be metal-independent. The active center of the enzyme contains the carboxylic group of the C-terminal aminoacid and imidazole group of histidine.


Keywords


α-galactosidase, Aspergillus niger, physicochemical properties, kinetic parameters, active center

Full Text:

PDF

References


1. Albershein P., Nevis D.J., English P.D., Karr A. A method of analysis of sugars in plant cell wall polysaccharides by gas-liquid chromatography. Carbohydr. Res, 1976; 5(3): 340-345.
https://doi.org/10.1016/S0008-6215(00)80510-8

2. Ananichev A.V., Ulezlo E.V., Zaprometova O.M., Bezborodov A.M. Some properties of immobilizated α-galactosidase from Cephalosporium acremonium 237. Applied Chemistry and Microbiology, 1987; 23(4): 552-557.

3. Andrews P. Estimation of the molecular weights of proteins by Sephadex gel-filtration. Biochem. J, 1964; 91(2): 222-233.
https://doi.org/10.1042/bj0910222
PMid:4158310 PMCid:PMC1202876

4. Buglova T.T., Malanchuk V.M., Zakharova I.Ya., e. a. Isolation, purification and some properties of fungal α-galactosidase. Microbiological Journal (Ukrainian), 1995; 57(1): 43-48.

5. Chaplin M.E., Kennedy J.E. Carbohydrate Analysis. Oxford; Washington: IRL Press, 1986. 228 p.

6. Civas A., Eberhard R., Dizet P., Ze Petek F. Glycosidases induced in Aspergillus tamarii. Selected α-D-galactosidase and α-mannanase. Biochem. J, 1984; 219(3): 857-863.
https://doi.org/10.1042/bj2190857
PMid:6331399 PMCid:PMC1153554

7. Dey D.F. Malhotra O.P. Kinetic behavior of sweet almound α-galactosidase. Biochem. Biophys. Acta, 1965; 185 (2): 402-408.
https://doi.org/10.1016/0005-2744(69)90433-1

8. Dey D.F., Pridham J. B., Sumar N. Multiple forms of Vicia faba α-galactosidases and their relationships. Phytochemistry, 1982; 21(9): 2195-2199.
https://doi.org/10.1016/0031-9422(82)85176-5

9. Dixon M., Webb L. Enzyme. Moscow: Mir, 1982. 235 p.

10. Dubois M., Gilles K.A., Hamilton J.K., e. a. Colorimeric method for determination of sugars and related substances. Anal. Chem, 1956; 8 (3): 350-356.
https://doi.org/10.1021/ac60111a017

11. Edsall J., Neuruth H., Walley K., e. a. Proteins. New-York: Acad. Press, 1958. 990 p.

12. Kornish-Bouden E. The Basis of Enzymatic Kinetics. Moscow: Mir, 1979. 280 p.

13. Kubo S. Glycosidases from soil microorganisms. J. Forensic Sci, 1989; 34(1): 96-104.
https://doi.org/10.1520/JFS12615J

14. Kurganov B.I., Petushkova E.B. Analysis of bell-like pH-dependences of enzyme reaction rate. Biochemistry, 1992; 57(3): 348-361.

15. Laemmli U.K. Cleavage of proteins during the assembly of the head of bacteriophage T4. Nature, 1970; 227: 680-685.
https://doi.org/10.1038/227680a0
PMid:5432063

16. Lowry O.H., Rosebrough N.J., Farr A.L, Randall R.J. Protein measurement with folinphenol reagent. J. Biol. Chem, 1951; 193(2): 265-275.

17. Neustroev K.N., Krilov A.S., Abroskina O.N., e. a. Isolation and properties of α-galactosidase from Aspergillus awamori. Biochemistry, 1991; 56(3): 447-457.

18. Petrova I.S., Vinzyunajte M.H. Applied Biochemistry and Microbiology, 1966; 2(1): 322-327.

19. Severin S. E., Solovyeva G.A. Manuals in Biochemistry. MSU, 1989. 509 p.

20. Somogy M. Notes on Sugar Determination. J. Biol. Chem, 1952; 195(1): 19-23.

21. Ulezlo E.V., Zaprometova O.M. α-Galactosidase of microorganisms. Applied Biochemistry and Microbiology, 1982; 18(1): 3-15.

22. Vidershine G.Ya. Biochemical Fundamentals of Glycosides. Moscow: Mir, 1980. 239 p.

23. Yan Z., Zhu Q., Shu M., e. a. Purification and properties of α-galactosidase from Absidia ramose. Acta Microbiol. Sin, 1989; 29(4): 265-271.


Refbacks

  • There are currently no refbacks.


Copyright (c) 2007 Studia biologica

Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 International License.