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From 2013.igem.org

Abstract

The human peptide LL-37 is a cationic antimicrobial peptide with activity against Gram-negative and Gram-positive bacteria. It has been shown to protect against Helicobacter pylori infection and its related gastritis, besides of inhibiting the growth of other pathogenic bacteria in the gastrointestinal tract such as Salmonella typhimurium and Escherichia coli. Most of the current synthetic expression systems for LL-37 depend on the construction of soluble fusion partners to avoid cytotoxic effects of the antimicrobial peptide in the E. coli host strain. However, these fusion systems require additional cleavage steps using enzymatic or chemical methods, which makes them impossible to express an active LL-37 peptide in vivo. In order to create a resistant host that can export LL-37 to the media, we intend to overexpress the E. coli acrAB and tolC operons, which activate the AcrAB-TolC efflux pump, a mechanism that has been related with resistance to this and similar antimicrobial peptides by expulsion. Our aim is to create a system in which E. coli expels LL-37 only when H. pylori or other pathogenic bacteria, such as S. typhimurium, E. coli or V. cholerae, are present. In order to do this we are using the LsrA promoter, which allows transcription in the presence of AI-2, a molecule produced by bacteria to communicate via quorum-sensing. E. coli also produces AI-2, so we designed an antisense RNA with specific secondary structure to inhibit the translation of LuxS, the enzyme responsible of the production of AI-2 in E. coli.