Figure 1

Characterization of aDec-expressing L. plantarum strains. a Schematic representation of the three anchors used. b Expression cassettes in which the gene fragment encoding the desired protein is translationally fused to the inducible P _sppA promoter [47]. The cassette was PCR-generated using the primers listed in Table 2 and inserted into previously described anchoring vectors [31, 36] digested with the restriction enzymes indicated in the Figure, as summarized in Table 3. All constructs include a N-terminal signal sequence (SP) for secretion and a HA-tag for immune detection. Three different anchoring domains were used, as described in the main text. Note that in the CWA construct, the anchoring domain is located C-terminally, meaning that the HA-tag will protrude from the cells after secretion and subsequent anchoring. c Growth curves for Lp-WT (control strain) and aDec-expressing strains; protein production was induced by addition of peptide pheromone at OD600 ~0.3. d Western blot analysis of cell-free protein extracts from aDec-expressing strains harvested 2 h after induction, using a mouse anti-HA primary antibody and a HRP-conjugated goat anti-mouse secondary antibody. e Flow cytometry analysis of the presence of aDec at the surface of Lp-WT (in black) compared to Lp-Lip-aDec, Lp-CWA-aDec or Lp-LysM-aDec (all in red).