SYNTHESIS AND MATURATION OF GREEN FLUORESCENT PROTEIN IN A CELL-FREE TRANSLATION SYSTEM

A cell-free translation system producing mature green fluorescent prot ein (GFP) can be a useful tool for studying the mechanism and kinetics of GFP chromophore formation, as well as for fast protein engineering . We report here that the mature GFP can be formed in the cell-free tr anslation system from E. coli. The synthesis of GFP in the cell-free s ystem reaches a plateau in 30 to 40 min whereas its maturation is comp leted in 4 h from the beginning of translation. The delay between the GFP synthesis and the chromophore formation in the cell-free system pr ovides the possibility to isolate and to analyse maturation intermedia tes for elucidation of the modification pathway.