The thermophilic fungus Humicola sp constitutively produces intracellular α-galactosidase (1.33 U mg−1 protein) within 48 h at 45°C in shaken flasks, when grown in a medium containing 7% wheat bran extract as a carbon source and 0.5% yeast extract as a nitrogen source. The enzyme has been purified to homogeneity by ultrafiltration, ethanol precipitation, DEAE cellulose and Sephacryl S-300 chromatography with a 124-fold increase in specific activity and 29.5% recovery. The molecular weight of the enzyme is 371.5 kDa by gel filtration on Sephacryl S-300 and 87.1 kDa by SDS-polyacrylamide gel electrophoresis. The enzyme has an optimum temperature of 65°C and an optimum pH of 5.0. Humicola α-galactosidase is a glycoprotein with 8.3% carbohydrate content and is acidic in nature with a pI of 4.0. The K m S for p-nitrophenyl-α-D-galactopyranoside, O-nitrophenyl-α-D-galactopyranoside, raffinose and stachyose are 0.279, 0.40, 1.45 and 1.42 mM respectively. The enzyme activity was strongly inhibited by Ag+ and Hg2+. D-Galactose inhibited α-galactosidase competitively and the inhibition constant (K i) for galactose was 11 mM.
Similar content being viewed by others
Author information
Authors and Affiliations
Additional information
Received 28 January 1999/ Accepted in revised form 07 April 1999
Rights and permissions
About this article
Cite this article
Kotwal, S., Gote, M., Khan, M. et al. Production, purification and characterization of a constitutive intracellular α-galactosidase from the thermophilic fungus Humicola sp. J Ind Microbiol Biotech 23, 661–667 (1999). https://doi.org/10.1038/sj.jim.2900680
Issue Date:
DOI: https://doi.org/10.1038/sj.jim.2900680