Identification of extracellular surface-layer associated proteins in Lactobacillus acidophilus NCFM

Abstract

Bacterial surface (S-) layers are crystalline arrays of self-assembling, proteinaceous subunits called S-layer proteins (Slps), with molecular masses ranging from 40 to 200 kDa. The S-layer-forming bacterium Lactobacillus acidophilus NCFM expresses three major Slps: SlpA (46 kDa), SlpB (47 kDa) and SlpX (51 kDa). SlpA has a demonstrated role in adhesion to Caco-2 intestinal epithelial cells in vitro, and has been shown to modulate dendritic cell (DC) and T-cell functionalities with murine DCs. In this study, a modification of a standard lithium chloride S-layer extraction revealed 37 proteins were solubilized from the S-layer wash fraction. Of these, 30 have predicted cleavage sites for secretion, 24 are predicted to be extracellular, six are lipid-anchored, three have N-terminal hydrophobic membrane spanning regions and four are intracellular, potentially moonlighting proteins. Some of these proteins, designated S-layer associated proteins (SLAPs), may be loosely associated with or embedded within the bacterial S-layer complex. Lba-1029, a putative SLAP gene, was deleted from the chromosome of L. acidophilus. Phenotypic characterization of the deletion mutant demonstrated that the SLAP LBA1029 contributes to a pro-inflammatory TNF-α response from murine DCs. This study identified extracellular proteins and putative SLAPs of L. acidophilus NCFM using LC-MS/MS. SLAPs appear to impart important surface display features and immunological properties to microbes that are coated by S-layers.

Identification of proteins involved in the anti-inflammatory properties of Propionibacterium freudenreichii by means of a multi-strain study

“…During this work, we identified the main actors explaining this phenotype. Thus, as has been demonstrated for lactobacilli and bifidobacteria2634, the immunomodulatory response is the result of the detection by immunocompetent cells of a mosaic of surface proteins and other components (pro-and anti-inflammatory proteins). All these components contribute to the net balance of immunomodulatory properties of Pf strains.…”

Murein hydrolase activity of surface layer proteins from Lactobacillus acidophilus against Escherichia coli

“…The SLPs of L. acidophilus fb11 also contributed positively to the probiotic activities of strains (data not shown). Johnson et al [4] identified 37 surface-layer associated proteins from L. acidophilus NCFM by lithium chloride extraction, including three major SLPs: SlpA (46 kDa), SlpB (47 kDa) and SlpX (51 kDa). In this study, the major lytic band was 46 kDa.…”

“…Many species of the genus Lactobacillus have been found to possess SLPs [3] and the structures and functions of those of L. acidophilus strains have been investigated extensively. Johnson et al [4] reported a modified method to extract the surface-layer associated proteins of L. acidophilus NCFM using lithium chloride that revealed 37 proteins, some of which might be loosely associated with or embedded within the bacterial surface layer complex and appeared to contribute considerably to the immunological properties and surface display features of the microbes.…”

Surface proteins of Propionibacterium freudenreichii are involved in its anti-inflammatory properties

“…None of the 3 proteins, previously thought to be composed of a 630 single protein molecular species, are more complex in631 Gram-positive bacteria such as L. acidophilus, the S-layer of 632 which contains 37 proteins[61], in addition to the previously 633 described SlpA. In P. freudenreichii ITG P20, at least 5 proteins634 are extracted using guanidine, SlpB being the major one, with 635 a MS-measured mass of 54,147 Da, which is close to the 636 54,145 Da size predicted for SlpB after cleavage of the signal 637 peptide (this work, predicted from the slpB gene).…”