Optimized growth kinetics of Pichia pastoris and recombinant Candida rugosa LIP1 production by RSM.
A predictive model for Pichia pastoris expression of highly active recombinant Candida rugosa LIP1 was developed by combining the Gompertz function and response surface methodology (RSM) to evaluate the effect of yeast extract concentration, glucose concentration, temperature, and pH on specific responses. Each of the responses (maximum population densities, specific growth rate (mumax), protein concentration, and minimum lag phase duration) was determined using the modified Gompertz function. RSM and 4-factor-5-level central composite rotatable design (CCRD) were adopted to evaluate the effects of growth parameters, such as temperature (21.6-38.4 degrees C), glucose concentration (0.3-3.7%), yeast extract (0.16-1.84%), and pH (5.3-8.7) on the responses of P. pastoris growth kinetics. Based on ridge maximum analysis, the optimum population density conditions were: temperature 24.4 degrees C, glucose concentration 2.0%, yeast extract 1.5%, and pH 7.6. The optimum specific growth rate conditions were: temperature 28.9 degrees C, glucose concentration 2.0%, yeast extract 1.1%, and pH 6.9. The optimum protein concentration conditions were: temperature 24.2 degrees C, glucose concentration 1.9%, yeast extract 1.5%, and pH 7.6. Based on ridge minimum analysis, the minimal lag phase conditions were: temperature 32.3 degrees C, glucose concentration 2.1%, yeast extract 1.1%, and pH 5.4. For the predicted value, the maximum population density, specific growth rate, protein concentration, and minimum lag phase duration were 15.7 mg/ml, 3.4 h(-1), 0.78 mg/ml, and 4.2 h, and the actual values were 14.3 +/- 3.5 mg/ml, 3.6 +/- 0.6 h(-1), 0.72 +/- 0.2 mg/ml, and 4.4 +/- 1.6 h, respectively.
Chang
Shu-Wei
SW
Institute of Bioscience and Biotechnology, National Taiwan Ocean University, Keelung, Taiwan.
Shieh
Chwen-Jen
CJ
Lee
Guan-Chiun
GC
Akoh
Casimir C
CC
Shaw
Jei-Fu
JF
D002175:Candida / Q000201:enzymology*; D003470:Culture Media; D005285:Fermentation; D005656:Fungal Proteins / Q000096:biosynthesis* / Q000235:genetics; D005947:Glucose / Q000378:metabolism; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008049:Lipase / Q000096:biosynthesis* / Q000235:genetics; D010843:Pichia / Q000254:growth & development* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D013696:Temperature
D003470:Culture Media; D005656:Fungal Proteins; D011994:Recombinant Proteins; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D008049:Lipase; D005947:Glucose