ABSTRACT
Abstract This study was carried out to express human epidermal growth factor (hEGF) in Pichia pastoris GS115. For this aim, the hEGF gene was cloned into the pPIC9K expression vector, and then integrated into P. pastoris by electroporation. ELISA-based assay showed that the amount of hEGF secreted into the medium can be affected by the fermentation conditions especially by culture medium, pH and temperature. The best medium for the optimal hEGF production was BMMY buffered at a pH range of 6.0 and 7.0. The highest amount of hEGF with an average yield of 2.27 µg/mL was obtained through an induction of the culture with 0.5% (v/v) methanol for 60 h. The artificial neural network (ANN) analysis revealed that changes in both pH and temperature significantly affected the hEGF production with the pH change had slightly higher impact on hEGF production than variations in the temperature.
Subject(s)
Humans , Pichia/metabolism , Recombinant Proteins/metabolism , Epidermal Growth Factor/metabolism , Pichia/genetics , Recombinant Proteins/genetics , Enzyme-Linked Immunosorbent Assay , Gene Expression , Cloning, Molecular , Culture Media/chemistry , Epidermal Growth Factor/genetics , Fermentation , Hydrogen-Ion ConcentrationABSTRACT
Background: Inulinase is a versatile enzyme from glycoside hydrolase family which targets the beta-2, 1 linkage of fructopolymers. In the present study, the effect of medium composition and culture conditions on inulinase production by Aspergillus niger ATCC 20611 was investigated in shake-flasks. Results: The highest extracellular inulinase (3199 U/ ml) was obtained in the presence of 25 percent (w/v) sucrose, 0.5 percent (w/v) meat extract, 1.5 percent (w/v) NaNO3 and 2.5 mM (v/v) Zn2+, at initial pH of 6.5, temperature 35ºC and 6 percent (v/v) of spores suspension in the agitation speed of 100 rpm. Surfactants showed an inhibitory effect on enzyme production. The optimum temperature for inulinase activity was found to be 50ºC. TLC analysis showed the presence of both exo- and endo-inulinase. Conclusion: Sucrose, Zn2+, and aeration were found to be the most effective elements in inulinase production by A. niger ATCC 20611. TLC analysis also showed that the crude enzyme contained both endo and exo-inulinases. The strain is suggested as a potential candidate for industrial enzymatic production of fructose from inulin.
Subject(s)
Aspergillus niger/metabolism , Glycoside Hydrolases/biosynthesis , Culture Techniques , Fermentation , Hydrogen-Ion Concentration , TemperatureABSTRACT
This research emphasizes on single cell protein (SCP) production and Biochemical Oxygen Demand (BOD) removal from whey with mixed yeast culture. For this purpose, 11 yeast strains were isolated from dairy products (M1-M11) and the strains were identified by morphological and physiological properties. These yeast strains were tested for their ability to reduce the BOD and to produce SCP from whey. Among these strains, K. lactis (M2) had the most SCP production from whey with the yield of 11.79 g/l. Ammonium sulphate as nitrogen source had an increasing effect on biomass yield. The mixed culture of the isolated yeast strains with Saccharomyces cerevisiae was used in order to increase the biomass yield and BOD removal. The highest biomass yield (22.38 g/l) and reduction of initial BOD from 30000 to 3450 mg/l were obtained with the mixed culture of K. lactis (M2) and S. cerevisiae.