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Journal archive > 2006 > N 5 September-October


N. V. Borzova, L. D. Varbanets

Institute of Microbiology and Virology National Academy of Sciences of Ukraine, Kyiv;
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?-N-acetylgalactosaminidase has been isolated from liquid culture of micromycete Aspergillus niger and purified 600 times by ammonium sulphate precipitation followed by ion exchange and gel-filtration chromatography on TSK-gels with specific activity 10.5 U/mg of protein. The preparation was homogenic: its molecular mass by the data of gel-filtration on Sepharose 6B was 430 kDa, on PAAGE in the system of DDSNa – 70 kDa. That gives every reason to suppose oligomeric structure of the enzyme molecule. The carbohydrate component, including mannose, galactose, glucosamine and two nonidentified hexosamines was observed in ?-N-acetylgalactosaminidase. Thermo- and pH-optima were 60 °C and pH 3.5, respectively. The enzyme was thermo- and pH-stable, resistant in storage. The enzyme was found to exhibit strict specificity in respect of glycon. It was shown that enzyme was competitively inhibited by substrate and reaction product. Km and Vmax with respect to nitrophenyl substrate were 1.25 mM and 10.5 mkmole/min/mg of protein. The activity of glycosidase tested was independent of the presence of metal ions. The presence of carboxylic group of C-terminal aminoacid and imidazol group of hystidine in active centre of molecule was established. A number of natural and synthetic substrates were able to activate (50–200%) production of A. niger ?-N-acetylgalactosaminidase.

Key words: ?-N-acetylgalactosаminidase, Aspergillus niger, purification, physico-chemical properties, catalytic activity, active centre of enzyme.

The original article in Russian is available for download in PDF format.

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