ABSTRACT
The isopentenol utilization pathway (IUP) is potential in terpenoids
synthesis. This study aimed to construct IUP-employed E. colichassis for stably synthesizing terpenoids. As to effectiveness,
promotor engineering strategy was employed to regulate IUP expression
level, while ribosome-binding site (RBS) library of the key enzyme was
constructed for screening the optimal RBS, followed by optimization of
concentration of inducer and substrates, the titer of reporting
production, lycopene, from 0.087 to 8.67 mg/OD600. As
about stability, the IUP expression cassette was integrated into the
genome through transposition tool based on CRISPR-associated
transposases. Results showed that
the strain with 13 copies produced
1.78-fold lycopene titer that of
the controlled strain with IUP-harbored plasmid, and it exhibited stable
expression after ten successions while the plasmid loss was observed in
the controlled strain in the 3rd succession. This
strategy provides valuable information for rapid construction of highly
effective and stable chassis employing IUP for terpenoids production.