Transport-controlled growth decoupling for self-induced protein expression with a glycerol-repressible genetic circuit.

peer reviewedDecoupling cell formation from recombinant protein synthesis is a potent strategy to intensify bioprocesses. Escherichia coli strains with mutations in the glucose uptake components lack catabolite repression, display low growth rate, no overflow metabolism, and high recombinant protein yields. Fast growth rates were promoted by the simultaneous consumption of glucose and glycerol, and this was followed by a phase of slow growth, when only glucose remained in the medium. A glycerol-repressible genetic circuit was designed to autonomously induce recombinant protein expression. The engineered strain bearing the genetic circuit was cultured in 3.9 g L-1 glycerol + 18 g L-1 glucose in microbioreactors with online oxygen transfer rate monitoring. The growth was fast during the simultaneous consumption of both carbon sources (C-sources), while expression of the recombinant protein was low. When glycerol was depleted, the growth rate decreased, and the specific fluorescence reached values 17% higher than those obtained with a strong constitutive promoter. Despite the relatively high amount of C-source used, no oxygen limitation was observed. The proposed approach eliminates the need for the substrate feeding or inducers addition and is set as a simple batch culture while mimicking fed-batch performance

Quantitative Systems Biology for Engineering Organisms and Pathways

Studying organisms as a whole for potential metabolic(ally) engineering of organisms for production of (bio)chemicals is essential for industrial biotechnology. To this end, integrative analysis of different –omics measurements (transciptomics, proteomics, metabolomics, fluxomics) provides invaluable information. Combination of experimental top-down and bottom-up approaches with powerful analytical tools/techniques and mathematical modeling, namely (quantitative) systems biology, currently making the state of art of this discipline, is the only practice that would improve our understanding for the purpose. The use of high-throughput technologies induced the required development of many bioinformatics tools and mathematical methods for the integration of obtained data. Such research is significant since compiling information from different levels of a living system and connecting them is not an easy task. In particular, construction of dynamic models for product improvement has been one of the goals of many research groups. In this Research Topic, we summarize and bring a general review of the most recent and relevant contributions in quantitative systems biology applied in metabolic modeling perspective. We want to make special emphasis on the techniques that can be widely implemented in regular scientific laboratories and in those works that include theoretical presentations. With this Research Topic we discuss the importance of applying systems biology approaches for finding metabolic engineering targets for the efficient production of the desired biochemical integrating information from genomes and networks to industrial production. Examples and perspectives in the design of new industrially relevant chemicals, e.g. increased titer/productivity/yield of (bio)chemicals, are welcome. Addition to the founded examples, potential new techniques that would frontier the research will be part of this topic. The significance of multi ‘omics’ approaches to understand/uncover the pathogenesis/mechanisms of metabolic diseases is also one of the main topics

<i>Vitreoscilla</i> Haemoglobin: A Tool to Reduce Overflow Metabolism

Overflow metabolism is a phenomenon extended in nature, ranging from microbial to cancer cells. Accumulation of overflow metabolites pose a challenge for large-scale bioprocesses. Yet, the causes of overflow metabolism are not fully clarified. In this work, the underlying mechanisms, reasons and consequences of overflow metabolism in different organisms have been summarized. The reported effect of aerobic expression of Vitreoscilla haemoglobin (VHb) in different organisms are revised. The use of VHb to reduce overflow metabolism is proposed and studied through flux balance analysis in E. coli at a fixed maximum substrate and oxygen uptake rates. Simulations showed that the presence of VHb increases the growth rate, while decreasing acetate production, in line with the experimental measurements. Therefore, aerobic VHb expression is considered a potential tool to reduce overflow metabolism in cells