The search for Lactobacillus proteins that bind to host targets : a thesis presented in partial fulfilment of the requirements for the degree of Master of Science in Microbiology at Massey University, Turitea, New Zealand

Abstract

Interactions between microorganisms and host cells in the gastrointestinal tract are crucial to the host’s health. Probiotic bacteria, such as the lactobacilli provide numerous benefits to human health thought to be mediated by bacterial proteins called effectors. Lactobacillus rhamnosus HN001 (L. rhamnosus HN001) is a cheesefermenting isolate with probiotic characteristics and Lactobacillus reuteri 100-23 (L. reuteri 100-23) is a coloniser of the rodent forestomach. Whereas L. rhamnosus HN001 was shown to reduce eczema in children, L. reuteri 100-23 reduces inflammation in mice. The effector proteins for these strains are largely unknown. In this thesis, phage display technology was used to search for proteins that bind specific ligands. Shot-gun genomic phage display library of L. rhamnosus HN001 was affinity screened on fibronectin as bait, leading to enrichment of specific recombinant clones. Analysis of 10 candidate clones, however, determined that these are not genuine binders, but may have been selected due to a potential growth advantage during amplification steps of the library. The L. reuteri 100-23 genomic shot-gun phage display library was subjected to two affinity screens on two baits: fibronectin and murine stomach tissue. The aim of the screen on the murine stomach tissue was to identify keratin-binding proteins, as this strain naturally colonises the murine keratinous forestomach. Whereas no enrichment was detected in the screen on fibronectin as a bait, a strong enrichment of a phagemid displaying a short peptide, IGINS, derived from a cell-surface protease of L. reuteri 100-23 was identified. Identifying and characterising probiotic bacterial proteins that positively influence health will lead to a greater understanding of gastrointestinal tract interactions. Ultimately, this aids development of probiotic use as therapeutic agents.