원문정보
초록
영어
Use of glucose as an energy source for ATP regeneration greatly improves the economical feasibility of cell-free protein synthesis. However, the protocols employing glucose, which was originally developed by Calhoun and Swartz, shows lower level of protein expression compared to other methods using the conventional energy sources such as creatine phosphate and
phosphoenolpyruvate. We found that the inefficiency of the glucose-utilizing cell-free protein synthesis was due to the accumulation of proton during the incubation periods. The productivity of target proteins was substantially enhanced simply by adjusting the pH of reaction mixture during the reactions of cell-free protein synthesis. When the pH of the reaction mixture was kept
constant by repeated additions of KOH, final amount of the accumulated chloramphenicol acetyltransferase reached approximately 1.5 mg/ml after 3 hours of incubation. Our results demonstrate the possibility of using cell-free protein synthesis as a realistic alternative to the conventional in vivo expression technologies.