PURIFICATION AND CHARACTERIZATION OF PULLULANASE FROM Lactococcus lactis |
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Authors: | Adam Waśko Zdzisław Targoński |
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Institution: | Department of Biotechnology, Human Nutrition and Science of Food Commodities , University of Life Sciences in Lublin, Poland , Lublin, Poland |
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Abstract: | This paper describes a simple and efficient method of isolation of a plullulanase type I from amylolytic lactic acid bacteria (ALAB). Extracellular pullulanase type I was purified from a cell-free culture supernatant of Lactococcus lactis IBB 500 by using ammonium sulfate fractionation and dialysis (instead of ultrafiltration), and ion-exchange chromatography with CM Sepharose FF followed by gel filtration chromatography with Sephadex G-150 as the final step. A final purification factor of 14.36 was achieved. The molecular mass of the enzyme was estimated as 73.9 kD. The optimum temperature for the enzyme activity was 45°C and the optimum pH was 4.5. Pullulanase activity was increased by addition Co2+ and completely inhibited by Hg2+. The enzyme activity was specifically directed toward α-1,6 glycosidic linkages of pullulan giving maltotriose units. Enzymatic hydrolysis of starch and amylose produced a mixture of maltose and maltotriose. |
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Keywords: | amylolytic lactic acid bacteria Lactococcus lactis pullulanase type I |
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