Fig. 1
Construction of nisin inducible rL. lactis vector expressing Hcp protein. A Schematic of NICE cassette of pNZ8048-spUSP45-hcp-CWAM6 plasmid under PnisA promoter for a surface-anchored form of Hcp. Colony PCR of positive transformant of rL. lactis cells harbouring hcp gene showing amplification at the expected size (~ 529 bp) (a). Schematic of rL. lactis having pSEC-spUSP45-hcp plasmid construct without CWAM6 cell wall anchor motif. Colony PCR of positive transformant of rL. Lactis harbouring hcp gene insert showing the amplified product at the expected size (~ 529 bp) (b). B Graphical depiction of surface display of Hcp protein anchored to the cell wall of rL. lactis via cell wall anchoring motif of M6 (CWAM6) protein of Streptococcus pyogenes (a). Graphical depiction of rL. lactis secreting Hcp in the extracellular matrix (b). Comparative growth profile of rL. lactis harbouring pNZ8048-spUSP45-hcp-CWAM6 and pSEC-spUSP45-hcp plasmid showing altered (reduced) growth kinetics of nisin-induced rL. lactis compared to empty L. lactis (NZ9000) and un-induced rL. lactis cells. Arrow indicates the time of nisin induction (c)