Abstract
Health-promoting aspects attributed to probiotic microorganisms, including adhesion to intestinal epithelia and modulation of the host mucosal immune system, are mediated by proteins found on the bacterial cell surface. Notably, certain probiotic and commensal bacteria contain a surface (S-) layer as the outermost stratum of the cell wall. S-layers are non-covalently bound semi-porous, crystalline arrays of self-assembling, proteinaceous subunits called S-layer proteins (SLPs). Recent evidence has shown that multiple proteins are non-covalently co-localized within the S-layer, designated S-layer associated proteins (SLAPs). In Lactobacillus acidophilus NCFM, SLP and SLAPs have been implicated in both mucosal immunomodulation and adhesion to the host intestinal epithelium. In this study, a S-layer associated serine protease homolog, PrtX (prtX, lba1578), was deleted from the chromosome of L. acidophilus NCFM. Compared to the parent strain, the PrtX-deficient strain (ΔprtX) demonstrated increased autoaggregation, an altered cellular morphology, and pleiotropic increases in adhesion to mucin and fibronectin, in vitro. Furthermore, ΔprtX demonstrated increased in vitro immune stimulation of IL-6, IL-12, and IL-10 compared to wild-type, when exposed to mouse dendritic cells. Finally, in vivo colonization of germ-free mice with ΔprtX led to an increase in epithelial barrier integrity. The absence of PrtX within the exoproteome of a ΔprtX strain caused morphological changes, resulting in a pleiotropic increase of the organisms’ immunomodulatory properties and interactions with some intestinal epithelial cell components.
Highlights
Lactic acid bacteria (LAB) are a clade of diverse Gram-positive, microaerophilic, and non-sporulating microbes which ferment hexoses primarily to lactic acid
One of the most prevalent S-layer associated proteins (SLAPs) in the non-covalently bound exoproteome of L. acidophilus NCFM is a 72 kDa uncharacterized serine protease encoded by the gene lba1578 (Johnson et al, 2013)
Because PrtX is an uncharacterized serine protease which is prominently featured in the non-covalent exoproteome in L. acidophilus, it was selected for functional analysis
Summary
Lactic acid bacteria (LAB) are a clade of diverse Gram-positive, microaerophilic, and non-sporulating microbes which ferment hexoses primarily to lactic acid Many of these bacteria, which include species from the genera Lactococcus, Enterococcus, Pediococcus, Oenococcus, Streptococcus, Leuconostoc, and Lactobacillus, have evolved through 1000s of years of fermentation in numerous food and drink substrates (Makarova et al, 2006; Douglas and Klaenhammer, 2010; Johnson and Klaenhammer, 2014). L. acidophilus NCFM is one of the most studied and well characterized probiotic bacteria (Sanders and Klaenhammer, 2001; Altermann et al, 2005; Klaenhammer et al, 2005, 2008)
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