Roberto Olivieri

Maximization of recombinant Helicobacter pylori neutrophil activating protein production in Escherichia coli: improvement of a chemically defined medium using response surface methodology.

Medium optimization for the production of constitutive recombinant Helicobacter pylori neutrophil activating protein (NAP) in Escherichia coli was investigated by using response surface methodology. Carbon to nitrogen ratio, concentrations of sodium polyphosphate and magnesium sulfate were considered as independent variables. The optimized medium was a chemically defined medium with a carbon to nitrogen ratio of 14.4 and with concentrations of sodium polyphosphate and magnesium sulfate about 7.1 g l(-1) and 3.04 g l(-1) respectively. The maximum recombinant NAP production level (1184.6 mg l(-1)) was 29.96% higher than that in control medium.

Studies on the maximization of recombinant Helicobacter pylori neutrophil-activating protein production in Escherichia coli: application of Taguchi robust design and response surface methodology for process optimization

The neutrophil-activating protein of Helicobacter pylori (HP-NAP) is a major antigen responsible for the generation of immune response in an infected individual. The cloning and expression of the gene corresponding to neutrophil-activating protein (NAP) were followed by process development for enhanced production and purification. The production process was developed in two parts. In the first part, some of the cultivation medium components (viz. carbon to nitrogen ratio, concentrations of sodium polyphosphate and magnesium sulphate) were optimized using the Taguchi robust experimental design. The intracellular NAP production level after 24 h of cultivation was considered as the target function or the dependent variable. There was a 76.8% increase in the NAP production level. Using this optimal medium composition obtained in the first part, the temperature of cultivation and the pH of cultivation medium were optimized in the second part. The NAP production level at hour 30 of cultivation was considered as the target function or the dependent variable. The optimal values for these two independent variables were 37.2 °C and 6.3 respectively. At this combination of temperature and pH, the theoretical maximum NAP production level was 1280 mg l−1. This optimal combination was verified experimentally and the NAP production level was found to be 1261 mg l−1. The optimization of the cultivation conditions resulted in a 61.5% increase in NAP production level. About a 2.91-fold overall increase in NAP production level at hour 24 of cultivation was achieved through process optimization.