Supporting Data for “Engineering of a Highly Efficient Escherichia coli Strain for Mevalonate Fermentation through Chromosomal Integration”

Full description in the file "WangReadme.rtf". The files below includes the pathway design (Fig. 1) and the bioengineering experimental data. Figure numbers in associated paper also include raw experimental data. The concentrations of feedstock (glucose), intermediate (acetate and pyruvate), and product (mevalonate) were measured by high-performance liquid chromatography (HPLC). The dataset includes one ChemDraw (.cdx) file. CDX files can be opened with proprietary ChemDraw 12.0, distributed by CambridgeSoft.These files contain data along with associated output from instrumentation supporting all results reported in Wang, J.; Niyompanich, S.; Tai, Y.-S.; Wang, J.; Bai, W.; Mahida, P.; Gao, T.; Zhang, K. Engineering of a highly efficient escherichia coli strain for mevalonate fermentation through chromosomal integration. Appl. Environ. Microbiol., 2016, 82, 7176–7184. Chromosomal integration of heterologous metabolic pathways is optimal for industrially relevant fermentation, as plasmid-based fermentation causes extra metabolic burden and genetic instabilities. In this work, chromosomal integration was adapted for the production of mevalonate, which can be readily converted into β-methyl-δ-valerolactone, a monomer for the production of mechanically tunable polyesters. The mevalonate pathway, driven by a constitutive promoter, was integrated into the chromosome of Escherichia coli to replace the native fermentation gene adhE or ldhA. The engineered strains (CMEV-1 and CMEV-2) did not require inducer or antibiotic and showed slightly higher maximal productivities (0.38 to ∼0.43 g/liter/h) and yields (67.8 to ∼71.4% of the maximum theoretical yield) than those of the plasmid-based fermentation. Since the glycolysis pathway is the first module for mevalonate synthesis, atpFH deletion was employed to improve the glycolytic rate and the production rate of mevalonate. Shake flask fermentation results showed that the deletion of atpFH in CMEV-1 resulted in a 2.1-fold increase in the maximum productivity. Furthermore, enhancement of the downstream pathway by integrating two copies of the mevalonate pathway genes into the chromosome further improved the mevalonate yield. Finally, our fed-batch fermentation showed that, with deletion of the atpFH and sucA genes and integration of two copies of the mevalonate pathway genes into the chromosome, the engineered strain CMEV-7 exhibited both high maximal productivity (∼1.01 g/liter/h) and high yield (86.1% of the maximum theoretical yield, 30 g/liter mevalonate from 61 g/liter glucose after 48 h in a shake flask).This research was supported by a grant from the National Science Foundation through the Center for Sustainable Polymers (grant CHE-1413862)