Metabolic responses to recombinant bioprocesses in Escherichia coli

Abstract

Escherichia coli has been widely used for the production of recombinant proteins. However, the unbalances between host metabolism and recombinant biosynthesis continue to hamper the efficiency of these recombinant bioprocesses. The additional drainage of biosynthetic precursors toward recombinant processes burdens severely the metabolism of cells that, ultimately, elicits a series of stress responses, reducing biomass growth and recombinant protein production. Several strategies to overcome these metabolic limitations have been implemented; however, in most cases, improvements in recombinant protein expression were achieved at the expense of biomass growth arrest, which significantly hampers the efficiency of recombinant bioprocesses. With the advent of high throughput techniques and modelling approaches that provide a system-level understanding of the cellular systems, it is now expected that new advances in recombinant bioprocesses are achieved. By providing means to deal with these systems, our understanding on the metabolic behaviour of recombinant cells will advance and can be further explored to the design of suitable hosts and more efficient and cost-effective bioprocesses. Here, we review the major metabolic responses associated with recombinant processes and the engineering strategies relevant to overcome these stresses. Moreover, the advantages of applying systems levels engineering strategies to enhance recombinant protein production in E. coli cells are discussed and future perspectives on the advances of mathematical modelling approaches to study these systems are exposed.

Statistical optimization of α-amylase production by Escherichia coliusing extruded bean as nitrogen and carbon source

“…Despite the culture medium containing extruded common bean had increased the growth of E. coli, changes in medium composition can interfere with the protein expression profile (Potvin et al, 2012;Carneiro et al, 2013). Aiming to verify the effectiveness of α-amylase production by E.colipAC92in different composition media, tests were performed and the results are demonstrated in Table 2.…”

“…In addition, due to rising prices of nitrogen and carbon sources, the bio-based enzymes production has been emerged as a promissory technology in biotechnology field. It is known that the development of a bio-based industrial enzymes production requires a low-cost medium and a versatile producing organism able to utilize a wide range of low-cost feedstock (Carneiro et al, 2013). In this scenario, hardened beans, an agroindustrial by-product that contains high amount of carbohydrates and proteins could be a promising nutrient source for α-amylase production.…”

“…However, the microbial production of amylases can be affected by certain factors, including microbial strain, culture medium formula and physicochemical conditions (Hii et al, 2012). Considering the composition of culture medium, the nature of nitrogen and carbon source could influence the rate of amylase production and, therefore the enhancement of target enzyme productivity in any fermentation system could be achieved through the improvement of the culture medium composition (Ye et al, 2010;Hortsch and Weuster-Botz, 2011;Boumba et al, 2013;Carneiro et al, 2013). Conventional change one-factor-at-a-time approach has been applied to optimize the enzyme production using different medium constituents.…”

“…However, this technique is time-consuming, expensive, and often leads to misinterpretationof results, once the interactions among parametersare not taken into account (Hii et al, 2012). An alternative to overcome this technical limitation is the use of statistical designs coupled to response surface methodology, which allows the study of several factors influencing the responses by varying them simultaneously and carrying out a limited number of experiments (Ye et al, 2010;Carneiro et al, 2013). The objective of this study was to optimize the α-amylase production by Escherichia coli pAC92 using extruded bean as nitrogen and carbon source.…”

Biofuel production: an odyssey from metabolic engineering to fermentation scale-up

“…Current molecular biology techniques can effectively alter enzyme levels to increase the flux toward biofuel synthesis. Common strategies include choice of plasmids and its copy numbers, promoter engineering, codon optimization, synthetic scaffolds, directed evolution/modification of key enzymes, improvement of ribosome binding sites, and knockout/knockdown of competitive pathways (Dueber et al, 2009; Carneiro et al, 2013; Nowroozi et al, 2014). New genetic techniques, such as RNA Interference, CRISPRs or TALENs, offer new capabilities to edit microbial metabolisms (Pratt and MacRae, 2009; Jiang et al, 2013; Sun and Zhao, 2013).…”