title: Purification and characterization of pullulanase from Lactococcus lactis.
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 Co(2+) and completely inhibited by Hg(2+). 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.
authors.affiliation: Department of Biotechnology, Human Nutrition and Science of Food Commodities, University of Life Sciences in Lublin, Poland.
authors.forename: Adam
authors.initials: A
authors.lastname: Waśko
authors.forename: Magdalena
authors.initials: M
authors.lastname: Polak-Berecka
authors.forename: Zdzisław
authors.initials: Z
authors.lastname: Targonski
mesh_terms: D001426:Bacterial Proteins; D005591:Chemical Fractionation / Q000379:methods*; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange / Q000379:methods*; D004795:Enzyme Stability; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D013294:Lactococcus lactis / Q000201:enzymology* / Q000378:metabolism; D008970:Molecular Weight; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D001426:Bacterial Proteins; D006026:Glycoside Hydrolases; C100181:pullulanase