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Purification and Characterization of Thermostable α-Galactosidase from Aspergillus terreus GR

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Abstract

An extracellular thermostable α-galactosidase producing Aspergillus terreus GR strain was isolated from soil sample using guar gum as sole source of carbon. It was purified to apparent homogeneity by acetone precipitation, gel filtration followed by DEAE-Sephacel chromatographic step. The purified enzyme showed a single band after sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The molecular weight of the purified enzyme after SDS-PAGE was 108 kDa. The enzyme showed optimum pH and temperature of 5.0 and 65 °C, respectively, for artificial substrate pNPαGal. α-Galactosidase from A. terreus GR is found to be thermostable, as it was not inactivated after heating at 65 °C for 40 min. The K m for pNPαGal, oNPαGal, raffinose, and stachyose are 0.1, 0.28, 0.42, and 0.33 mM, respectively. Inhibitors such as 1,10-phenanthroline, phenylmethylsulfonyl fluoride, ethylenediaminetetraacetic acid, mercaptoethanol, and urea have no effect, whereas N-bromosuccinamide inhibited enzyme activity by 100%. Among metal ions tested, Mg2+, Ni2+, Ca2+, Co2+, and Mn2+ had no effect on enzyme activity, but Ag+, Hg2+, and Cu2+ have inhibited complete activity.

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Acknowledgment

One of the authors, Shankar S. K., thanks Council of Scientific and Industrial Research (CSIR) New Delhi, India for providing financial assistance in the form of Senior Research Fellowship (SRF) during this work.

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  1. Department of Biochemistry, Gulbarga University, Gulbarga, 585106, Karnataka, India

    S. K. Shankar, S. K. Dhananjay & V. H. Mulimani

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  1. S. K. Shankar
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  2. S. K. Dhananjay
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  3. V. H. Mulimani
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Correspondence to V. H. Mulimani.

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Shankar, S.K., Dhananjay, S.K. & Mulimani, V.H. Purification and Characterization of Thermostable α-Galactosidase from Aspergillus terreus GR . Appl Biochem Biotechnol 152, 275–285 (2009). https://doi.org/10.1007/s12010-008-8271-7

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