Purification and characterization of a thermostable α-galactosidase from Thielavia terrestris NRRL 8126 in solid state fermentation

Saad, Rawia and Fawzi, Eman (2012) Purification and characterization of a thermostable α-galactosidase from Thielavia terrestris NRRL 8126 in solid state fermentation. Acta Biologica Hungarica, 63 (1). pp. 138-150. ISSN 0236-5383

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Abstract

Several seeds and husks of some plants belonging to leguminosae, Graminae, Compositae and Palmae were evaluated as carbon substrates to produce α-galactosidase (α-Gal) by the thermophilic fungus, Thielavia terrestris NRRL 8126 in solid substrate fermentation. The results showed that Cicer arietinum (chick pea seed) was the best substrate for α-Gal production. The crude enzyme was precipitated by ammonium sulphate (60%) and purified by gel filtration on sephadex G-₁₀₀ followed by ion exchange chromatography on DEAE-Cellulose. The final purification fold of the enzyme was 30.42. The temperature and pH optima of purified α-Gal from Thielavia terrestris were 70 °C and 6.5, respectively. The enzyme showed high thermal stability at 70 °C and 75 °C and the half-life of the α-Gal at 90 °C was 45 min. K_m of the purified enzyme was 1.31 mM. The purified enzyme was inhibited by Ag²⁺, Hg²⁺, Zn²⁺, Ba²⁺, Mg²⁺, Mn²⁺ and Fe²⁺ at 5 mM and 10 mM. Also, EDTA, sodium arsenate, L-cysteine and iodoacetate inhibited the enzyme activity. On the other hand, Ca²⁺, Cu²⁺, K⁺ and Na⁺ slightly enhanced the enzyme activity at 5 mM while at 10 mM they caused inhibition. The molecular weight of the α-Gal was estimated to be 82 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This enzyme displays a number of biochemical properties that make it a potentially strong candidate for biotechnological and medicinal applications.

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