A procedure to identify peptides with a-AMP activity

We showed previously that insertion of Synechocystis ?12-desaturase in Salmonella’s membrane alters membrane physical state (MPS) followed by the expression of stress genes causing inability to survive within murine macrophages . Recently, we showed that expression of one membrane lipid domain (MLD) of ?12-desaturase (ORF200) interferes with Salmonella MPS causing loss of virulence in mice and immunoprotection. We postulated that a putative a-AMP intercalates specifically within phospholipids but, depending on its amino acid sequence, does so within particular key sensors of MLD. In this study we choose as target for a putative synthetic AMP, PhoP/PhoQ, a sensor that responds to low Mg2+ concentration during murine macrophages infection. We synthesized a modified DNA fragment coding for an amino acid sequence (NUF) similar to that fragment and expressed it in Salmonella LT2 strain. We show that the pattern of gene expression controlled by PhoP/PhoQ and other pathways involving phospholipids biosynthesis, stress proteins and genes coding for antigens are dysregulated. We also present RNAseq of strain expressing ORF200 and showed that the pattern of the same genes is altered. Accumulation of NUF conferred temporary immunoprotection suggesting that it is possible to address a synthetic peptide to a specific MLD. This represents a powerful procedure to synthesize a-AMPs and generate live non virulent strains for vaccination.