Purification and Characterization of Extracellular Inulinase from a Marine Yeast <Emphasis Type="Italic">Cryptococcus aureus</Emphasis> G7a and Inulin Hydrolysis by the Purified Inulinase |
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Authors: | Jun Sheng Zhenming Chi Fang Gong Jing Li |
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Institution: | (1) Unesco Chinese Center of Marine Biotechnology, Ocean University of China, Yushan Road, No. 5, Qingdao, China |
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Abstract: | The extracellular inulinase in the supernatant of the cell culture of the marine yeast Cryptococcus aureus G7a was purified to homogeneity with a 7.2-fold increase in specific inulinase activity compared to that in the supernatant
by ultrafiltration, concentration, gel filtration chromatography (Sephadex™ G-75), and anion exchange chromatography (DEAE
sepharose fast flow anion exchange). The molecular mass of the purified enzyme was estimated to be 60.0 kDa. The optimal pH
and temperature of the purified enzyme were 5.0 and 50 °C, respectively. The enzyme was activated by Ca2+, K+, Na+, Fe2+, and Zn2+. However, Mg2+, Hg2+, and Ag+ acted as inhibitors in decreasing the activity of the purified inulinase. The enzyme was strongly inhibited by phenylmethanesulphonyl
fluoride (PMSF), iodoacetic acid, EDTA, and 1,10-phenanthroline. The K
m and V
max values of the purified enzyme for inulin were 20.06 mg/ml and 0.0085 mg/min, respectively. A large amount of monosaccharides
were detected after the hydrolysis of inulin with the purified inulinase, indicating the purified inulinase had a high exoinulinase
activity. |
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Keywords: | Inulinase Marine yeasts Inulin Characterization Cryptococcus aureus G7a |
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