Tag Archives: Sheffield

Systems Analyses Reveal the Resilience of Escherichia coli Physiology during Accumulation and Export of the Nonnative Organic Acid Citramalate

Joseph Webb, Vicki Springthorpe, Luca Rossoni, David-Paul Minde, Swen Langer, Heather Walker, Amias Alstrom-Moore, Tony Larson, Kathryn Lilley, Graham Eastham, Gill Stephens, Gavin H. Thomas, David J. Kelly, Jeffrey Green

Citramalate is an attractive biotechnology target because it is a precursor of methylmethacrylate, which is used to manufacture Perspex and other high-value products. Engineered E. coli strains are able to produce high titers of citramalate, despite having to express a foreign enzyme and tolerate the presence of a nonnative biochemical. A systems analysis of the citramalate fermentation was undertaken to uncover the reasons underpinning its productivity. This showed that E. coli readily adjusts to the redirection of metabolic resources toward recombinant protein and citramalate production and suggests that E. coli is an excellent chassis for manufacturing similar small, polar, foreign molecules. Read more

Proteomic Profiling, Transcription Factor Modeling, and Genomics of Evolved Tolerant Strains Elucidate Mechanisms of Vanillin Toxicity in Escherichia coli

Calum A. Pattrick, Joseph P. Webb, Jeffrey Green, Roy R. Chaudhuri, Mark O. Collins, David J. Kelly

A particular problem for the biotechnological production of many of the valuable chemicals that we are now able to manufacture in bacterial cells is that these products often poison the cells producing them. Solutions to improve product yields or alleviate such toxicity using the techniques of modern molecular biology first require a detailed understanding of the mechanisms of product toxicity. Here we have studied the economically important flavor compound vanillin, an aromatic aldehyde that exerts significant toxic effects on bacterial cells. We used high-resolution protein abundance analysis as a starting point to determine which proteins are upregulated and which are downregulated by growth with vanillin, followed by gene expression and mutant studies to understand the mechanism of the response. In a second approach, we evolved bacterial strains with higher vanillin tolerance. Their genome sequences have yielded novel insights into vanillin tolerance that are complementary to the proteomics data set. Read more

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