The Efficient Production of a Model Designed Peptide Using Engineered E. coli BL21(DE3) for Sucrose Metabolism

Designed peptide surfactants (DPS) have commercial potential due to their advanced properties. Recombinant production in E. coli from renewable carbon has the capacity to manufacture DPS at competitive prices with the existing petrochemical route. Sucrose from sugarcane is the ideal feedstock for microbial fermentation due to its superior environmental performances. Designed peptide surfactants however, are currently produced with an inefficient process from complex medium and E. coli does not metabolize sucrose. Furthermore, the amino acid DPS composition creates an unbalanced precursor drain in the host organism.

In this work, a model designed surfactant was efficiently produced from sucrose. Sucrose metabolism was engineered on E. coli BL21(DE3) and the model peptide produced from minimal medium. Production in glucose and sucrose was compared and high product titer was obtained using fed-batch operations. Flux Balance Analysis (FBA) suggested metabolic limitations during batch production and the cultivation medium was engineered to increase specific productivity.

Experimental results demonstrated the suitability of sucrose as a feedstock for microbial fermentation; more than 6 g/L of the recombinant peptide were obtained from sucrose or glucose and specific productivity reached about 20% maximum of the biomass content. FBA underlined the role of Methionine and Histidine biosynthesis for product formation and cell growth. Supplementing medium with the former residue increased final cell density and supplementing the latter residue increased specific productivity to 85% of that of complex medium. To the best of our knowledge this is the first report of designed peptides manufacture using sucrose feedstock as well as the highest product titer documented to date.