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Members of the genus  Lactobacillus  have a long history in food applications and are considered as promising and safe hosts for delivery of medically interesting proteins. We have assessed multiple surface anchors derived from  Lactobacillus plantarum  for protein surface display in multiple  Lactobacillus  species, using a  Mycobacterium tuberculosis  hybrid antigen as test protein. The anchors tested were a lipoprotein anchor and two cell wall anchors, one non-covalent (LysM domain) and one covalent (sortase-based anchoring using the LPXTG motif). Thus, three different expression vectors for surface-anchoring were tested in eight  Lactobacillus  species. When using the LPXTG and LysM cell wall anchors, surface display, as assessed by flow cytometry and fluorescence microscopy, was observed in all species except  Lactobacillus acidophilus . Use of the cell membrane anchor revealed more variation in the apparent degree of surface-exposure among the various lactobacilli. Overproduction of the secreted and anchored antigen impaired bacterial growth rate to extents that varied among the lactobacilli and were dependent on the type of anchor. Overall, these results show that surface anchors derived from  L .  plantarum  are promising candidates for efficient anchoring of medically interesting proteins in other food grade  Lactobacillus  species.

Anchoring of heterologous proteins in multiple Lactobacillus species using anchors derived from Lactobacillus plantarum