A cell engineering approach to enzyme-based fed-batch fermentation

Abstract Background A fundamental problem associated with E. coli fermentations is the difficulty in achieving high cell densities batch cultures, attributed large part to production and accumulation of acetate through a phenomenon known as overflow metabolism when supplying enough glucose for density desired. Although fed-batch configuration standard method reducing such issues, traditional systems require components which become problematic applying them at smaller scale. One alternative has been development system whereby enzymatic degradation starch used release controlled rate. However, date, amylolytic enzymes have only applied culture exogenously, whereas our goal design construct self-secreting chassis capable self-regulated enzyme-based fermentation. Results putative glucoamylase from C. violaceum cloned expressed BL21(DE3) W3110, exhibits significant releasing activity. Extracellular activity was enhanced following replacement native signal peptide DsbA sequence, contributing secreting strain utilising sole carbon source defined media. Introduction PcstA , sensitive K12 compatible promoter, incorporation this alongside gave rise increased cultures grown on (OD 600 ∼ 30) compared those an equivalent amount 15). Lastly, novel fermentation demonstrated via simultaneous expression recombinant protein interest (eGFP), resulting fourfold increase yield media containing equivalent. Conclusions This study developed, secretion previously uncharacterised bacterial glucoamylase, direct conversion. The ability achieve well glucose, demonstrates first time engineering approach