Screening And Expression Of Protopectinase From Aspergillus Oryzae

Protopectinase is an enzyme that solubilizes protopectin forming highly polymerized soluble pectin. Aspergillus oryzae PO that showed maximum protopectinase activity was isolated from soil of persimmon orchard. The protopectinase production of Aspergillus oryzae PO was optimized in submerged fermentation using one-factor-at-a-time method and response surface methodology (RSM). Three fermentation variables (Temperature, NaN03and Apple pomace), exhibited greatest effects on protopectinase were chosen for RSM of Box-Behnken Design. Under the optimal fermentation conditions, the maximum protopectinase production indicated270.0U/ml.Polygalacturonase belonged to A-type protopectinase was cloned and expressed in Pichia pastoris GS115. The production level of polygalacturonase was0.418mg/ml. The specific activity of purified recombinant polygalacturonase was7519.54U/mg towards polygalacturonic acid. The optimal temperature and pH of recombinant polygalacturonase were45℃and5.0, respectively. The recombinant polygalacturonase activity was enhanced by the presence of Mg2+, while Ca2+, Ni2+, Mn2+, Cu2+and SDS strongly inhibited the enzyme activity. The apparent Km value and Vmax value were5.59mg/ml and1.01μmol (min·ml)-1.