Highly productive cell-free protein synthesis system using condensed wheat-germ extract

In order to increase the productivity of a cell-free protein synthesis system using wheat-germ extract, the extract was highly condensed by polyethylene glycol precipitation, which greatly increased both the final protein concentration and the initial synthesis rate of translation. However, the translational reaction using the condensed extract ceased earlier than the reaction using the uncondensed extract did, because ribonuclease and phosphatase activities, which destroyed mRNA and lessened the ATP level, respectively, were also condensed. Adding Cu2+ to the condensed translational system was found to reduce these activities resulting in a prolonged reaction with a high initial rate. Finally, 120 mu g of dihydrofolate reductase protein from Escherichia coli was obtained in 1 ml of reaction mixture in 3 h. In addition, the initial protein synthesis rate was 70 mu g ml(-1) h(-1), which was about 10 times higher than that of a conventional uncondensed extract.