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. 2009 Jun 26;75(16):5290–5299. doi: 10.1128/AEM.00621-09

FIG. 3.

FIG. 3.

Turbidimetric determination of PHB-degrading activities of PHB depolymerases. A decrease in the turbidity of a reaction mixture (200 μl) containing 0.7 mg of nPHB granules, aPHB granules, or dPHB was detected spectrophotometrically at 650 nm. (A to D) Three micrograms of purified His-tagged PhaZ1 from B. megaterium or His-tagged PhaZ from B. thuringiensis was used in each assay. nPHB granules were isolated from B. megaterium cells that were grown in LB medium supplemented with 2% sodium acetate for 6 h. Symbols: (A) solid circles, nPHB granules plus PhaZ1; solid triangles, nPHB granules plus PhaZ; hollow triangles, trypsin-pretreated nPHB granules plus PhaZ; hollow circles, nPHB granules alone; (B) solid circles, nPHB granules plus refolded PhaZ1; solid triangles, nPHB granules plus refolded PhaZ; hollow circles, nPHB granules alone; (C) solid circles, trypsin-pretreated nPHB granules plus refolded PhaZ1; solid triangles, trypsin-pretreated nPHB granules plus refolded PhaZ; hollow circles, trypsin-pretreated nPHB granules alone; (D) solid circles, aPHB granules plus PhaZ1; solid triangles, aPHB granules plus refolded PhaZ1; hollow circles, aPHB granules alone. (E and F) Different amounts of purified His-tagged PhaZ1 from B. megaterium or His-tagged PhaZ from B. thuringiensis were used in the assays. Symbols: (E) solid circles, dPHB plus 3 μg of PhaZ1; solid triangles, dPHB plus 3 μg of PhaZ; hollow circles, dPHB alone; (F) solid circles, dPHB plus 15 μg of PhaZ1; solid triangles, dPHB plus 15 μg of PhaZ; hollow circles, dPHB alone. OD650, optical density at 650 nm.