[논문]Purification and Characterization of Endo-<tex>$\\beta$</tex>-1,4 Mannanase from Aspergillus niger gr for Application in Food Processing Industry

Naganagouda, K.; Salimath, P.V.; Mulimani, V.H.

doi:10.4014/jmb.0901.029

Purification and Characterization of Endo-$\\beta$-1,4 Mannanase from Aspergillus niger gr for Application in Food Processing Industry 원문보기

Journal of microbiology and biotechnology, v.19 no.10, 2009년, pp.1184 - 1190

Naganagouda, K. (Department of Biochemistry, Gulbarga University) , Salimath, P.V. (Department of Biochemistry and Nutrition, Central Food Technological Research Institute) , Mulimani, V.H. (Department of Biochemistry, Gulbarga University)

Abstract ▼ AI-Helper

A thermostable extracellular $\beta$-mannanase from the culture supernatant of a fungus Aspergillus niger gr was purified to homogeneity. SDS-PAGE of the purified enzyme showed a single protein band of molecular mass 66 kDa. The $\beta$-mannanase exhibited optimum catalytic activity at pH 5.5 and $55^{\circ}C$. It was thermostable at $55^{\circ}C$, and retained 50% activity after 6 h at $55^{\circ}C$. The enzyme was stable at a pH range of 3.0 to 7.0. The metal ions $Hg^{2+}$, $Cu^{2+}$, and $Ag^{2+}$ inhibited complete enzyme activity. The inhibitors tested, EDTA, PMSF, and 1,10-phenanthroline, did not inhibit the enzyme activity. N-Bromosuccinimide completely inhibited enzyme activity. The relative substrate specificity of enzyme towards the various mannans is in the order of locust bean gum>guar gum>copra mannan, with $K_m$ of 0.11, 0.28, and 0.33 mg/ml, respectively. Since the enzyme is active over a wide range of pH and temperature, it could find potential use in the food-processing industry.

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9 ml of the substrate solution at 50℃ for 20 min. The amount of reducing sugars produced in the enzyme reaction was measured as D-mannose reducing equivalents by the 3, 5-dinitrosalicylic acid method [15].

성능/효과

0, 25 mM) at 10℃ for 12 h, with intermittent change of buffer. After a second step of purification (DEAE-Sephacel chromatography), 7.82-fdld of purification and overall yield of approximately 10% were observed (Table 1). The dialyzate from ion-exchange chromatography was concentrated to a small volume and subjected to gel filtration on a PD-10 column.
Kinetic parameters like and Vmax of Aspergillus niger gr β-mannanase are summarized in Table 2; the results revealed that β-mannanase had a higher affinity towards LBG (Ke, 0.11 mg/ml) than that _야f other mannans like guar gum and copra mannan. The relative substrate specificity of β-mannanase towards the various natural mannans was in the order of locust bean gum>guar gum>copra mannan (Table 2).
β-Mannanase secreted extracellularly by Aspergillus niger gr was purified to homogeneity by ammonium sulfate precipitation and ionexchange chromatography. The procedure yielded 1.46 mg of purified enzyme from 250 ml of culture filtrate, and the recovery of total β-mamanase activity was 9.5% (Table 1). After simple purification steps, PAGE and SDS-PAGE of the final enzyme preparation showed a sin_이e band.

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Purification and Characterization of Endo-$\\beta$-1,4 Mannanase from Aspergillus niger gr for Application in Food Processing Industry 원문보기

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Purification and Characterization of Endo-$\\beta$-1,4 Mannanase from Aspergillus niger gr for Application in Food Processing Industry 원문보기

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