Cooperativity of stabilized mRNA and enhanced translation activity in the cell-free system

Detailed analysis of cell-free translation, coupled transcription-translation in static conditions and a continuous flow system based on E. coli S30 extracts was performed. Degradation of template mRNA was the predominant trigger to terminate the protein synthesis. In a coupled system, mRNA was preserved by repeated transcription whereas the starvation of nucleotide triphosphates led to the termination of protein synthesis in less than 1 h. In the CFCF system, NTP was held at the level of initial concentration and therefore did not arrest the translation for 15 h. The accurate coupling of transcriptional rate and translational rate was also crucial to enhance the efficiency of protein synthesis.