3 June 2016

Redefining the Minimal Lipopolysaccharide Structure in Escherichia coli: Development of Endotoxin-free E.Coli Strains

Organised by:

SCI's Biotechnology Group in partnership with the University of Westminster

University of Westminster

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Recombinant proteins, commonly manufactured in Escherichia coli, are inherently contaminated with constituents of the bacterial cell such as lipopolysaccharide (LPS). LPS, also known as endotoxin, is a potent agonist for hTLR4/MD-2-mediated proinflammatory activity in human immune cells. To prevent toxicity, biological material prepared from E. coli must be essentially free of endotoxin.

However, efficient and cost-effective elimination of endotoxin is still a challenging task, and none of the downstream applications described thus far remove endotoxin entirely. As an alternative strategy for the production of endotoxin-free biomolecules, a set of non-conditional E. coli derivatives was constructed that lack all outer membrane agonists for hTLR4/MD-2 activation.

The mutant strains entirely lack LPS, yet remain viable despite exclusively elaborating the tetra-acylated, endotoxically inactive lipid A precursor lipid IVA. Viability requires a suppressor mutation that enables the strains to translocate lipid IVA across the inner membrane to support outer membrane biogenesis. Consistent with the results that activation of hTLR4/MD-2 signalling by the mutant cells was of several orders of magnitude lower compared with cells of the E. coli wild-type strain, heterologous proteins prepared from different mutant strains were free of endotoxic activity. This new series of genetically engineered E. coli strains lacking LPS holds promise for the manufacture of endotoxin-free products.


Day 1 - 03 June 2016

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University of Westminster

School of Life Sciences
115 New Cavendish Street
London W1W 6UW

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SCI Communications Team

Tel: +44 (0) 20 7598 1594

Email: communications@soci.org

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Dr Uwe Mamat, Division of Structural Biochemistry, Research Center Borstel, Leibniz-Center for Medicine and Biosciences