Novel surface display system for heterogonous proteins on Lactobacillus plantarum

Abstract

Aims: To establish a novel cell surface display system that would enable the display of target proteins on Lactobacillus plantarum. Methods and Results: BlastP analysis of the amino acids sequence data revealed that the N‐terminus of the putative muropeptidase MurO from L. plantarum contained two putative lysin motif (LysM) repeat regions, implying that the MurO was involved in bacterial cell wall binding. To investigate the potential of MurO for surface display, green fluorescent protein (GFP) was fused to MurO at its C‐terminus and the resulting fusion protein was expressed in Escherichia coli. After being mixed with L. plantarum cells in vitro, GFP was successfully displayed on the surfaces of L. plantarum cells. Increases in the fluorescence intensities of chemically pretreated L. plantarum cells compared to those of nonpretreated cells suggested that the peptidoglycan was the binding ligand for MurO. SDS sensitivity assay showed that the GFP fluorescence intensity was reduced after being treated with SDS. To demonstrate the applicability of the MurO‐mediated surface display system, β‐galactosidase from Bifidobacterium bifidium, in place of GFP, was functionally displayed on the surface of L. plantarum cells via MurO. Conclusions: The MurO was a novel anchor protein for constructing a surface display system for L. plantarum. Significance and Impact of Study: The success in surface display of GFP and β‐galactosidase opened up the feasibility of employing the cell wall anchor of MurO for surface display in L. plantarum.

Lactic Acid Bacteria as a Surface Display Platform for Campylobacter jejuni Antigens

“…It has been demonstrated that hybrid PA fusions exhibit similar properties [Bosma et al, 2006;Raha et al, 2005;Steen et al, 2003]. Also LysM domains of other Gram-positive bacteria bind heterologous proteins to the cell wall peptidoglycan [Hu et al, 2010;Turner et al, 2004;Xu et al, 2011].…”

Improvement of LysM-Mediated Surface Display of Designed Ankyrin Repeat Proteins (DARPins) in Recombinant and Nonrecombinant Strains of Lactococcus lactis and Lactobacillus Species

“…These include the bacteriophage endolysin Lyb5 (20) and the muropeptidase MurO from Lactobacillus plantarum (21). Differences in the ability to bind LysM repeat-containing proteins have been observed between Lactobacillus species, which reflect the differences in lactobacillus surfaces (22).…”

A genetically engineered protein domain binding to bacterial murein, archaeal pseudomurein, and fungal chitin cell wall material

“…Similarly, the PMB domains of archaeal hydrolases (PeiW and PeiP) allow binding of the enzymes to pseudomurein, facilitating hydrolysis by the catalytic domains (Luo et al 2002; Steenbakkers et al 2006; Visweswaran et al 2010). In recent years, the LysM domains have been widely applied for various biotechnological purposes such as in the preparation of oral influenza vaccine and for surface display of heterologous proteins on bacterial cell surfaces (Bosma et al 2006; Okano et al 2008; Shao et al 2009; Hu et al 2010; Saluja et al 2010; Xu et al 2011). Similarly, the PMB domain could be employed for the display of target proteins on pseudomurein-containing archaeal cell surfaces; it could also be used as a marker protein to identify pseudomurein-containing methanogens from their counterparts.…”