Fig 1.
Elution profile of Malus domestica (Golden delicious apple) fruit pulp PPO isoforms obtained from HPLC.
Superdex column was used at retention time (RT) 12.835 min with 0.3 absorbance on spectrophotometer. Time period (24 min) is plotted on horizontal axis and absorbance is plotted on vertical axis.
Table 1.
Stepwise purification result of the fruit pulp of Golden delicious apple polyphenol oxidase.
Fig 2.
SDS-PAGE gel electrophoresis of the purified polyphenol oxidase isoforms from the fruit pulp of Golden delicious apple.
Lane 1 & 2: molecular weight markers. Lane 3–5: apple PPO samples in triplicate. Gel is showing two active bands of PPO isoforms (28 and 45 kDa).
Fig 3.
pH optimization of polyphenol oxidase isoforms purified from the fruit pulp of Golden delicious apple.
Each bar plot is showing the mean value of the polyphenol oxidase activity at varying pH (3–8). Error bars are representation of standard error (SE) of the data. Horizontal axis is showing phosphate buffer of varying pH and vertical axis is showing polyphenol oxidase activity at different pH is plotted on vertical axis. Graph is showing maximum activity of Golden delicious apple fruit pulp PPO at pH 6 to 7.
Fig 4.
Optimization of temperature of the purified isoforms of polyphenol oxidase purified from the fruit pulp of Golden delicious apple.
Each bar plot is showing mean value of the polyphenol oxidase activity at varying temperatures (30°C-70°C). Error bars are representation of standard error (SE) of the data. Horizontal axis is showing the temperature in °C and vertical axis is showing the polyphenol oxidase activity. Graph is showing maximum activity of Golden delicious apple fruit pulp PPO isoforms at 40˚C.
Fig 5.
Effect of metal ions on the polyphenol oxidase activity of Golden delicious apple fruit pulp.
Each bar plot is showing the mean value of the polyphenol oxidase activity in the presence of metal ions. First bar represents mean value of the polyphenol oxidase activity in the absence of metal ions. The other three bar plots represent the polyphenol oxidase activity of apple fruit pulp in the presence of CuSO4, KCl, CaCl2. Error bars are representation of standard error (SE) of the data. Horizontal axis is showing the metal ion presence (1mM Conc) and vertical axis is showing polyphenol oxidase activity of the purified isoforms from Golden delicious apple fruit pulp.
Fig 6.
Lineweaver Burk plot for the determination of Km and Vmax of polyphenol oxidase.
Fig 7.
Graphs obtained from the applied biosystem’s pulse liquid protein sequencer.
(a-g) showing the result of N-terminal microsequences (AKITFHG) of purified polyphenol oxidase isoform (28 kDa) from the fruit pulp of Golden delicious apple.
Fig 8.
Graphs obtained from the applied biosystem’s pulse liquid protein sequencer.
(a-e) showing the N-terminal microsequences (APGGG) of the purified polyphenol oxidase isoform (45 kDa) from fruit pulp of Golden delicious apple fruit pulp.