P40 and P75 Are Singular Functional Muramidases Present in the Lactobacillus casei /paracasei/rhamnosus Taxon

Abstract

<i>Lactobacillus casei</i> and <i>Lactobacillus rhamnosus</i> proteins P40 and P75 belong to a large family of secreted cell wall proteins that contain a carboxy(C)-terminal CHAP or NlpC/P60 superfamily domains. In addition to their peptidoglycan hydrolases activity, proteins in this family are specific antigens of pathogens, frequently responsible of interactions with the host. <i>L. rhamnosus</i> GG and <i>L. casei</i> BL23 purified P40 and P75 proteins have antiapoptotic activity by inducing the EGF/Akt pathway. The aim of this work was to study the genetics, phylogeny and dissemination of this family of proteins in the genus <i>Lactobacillus</i> as well as their characteristics and likely function. The scrutiny of their DNA encoding sequences revealed the presence of minisatellite DNA in the P75 encoding gene of <i>L. casei/paracasei</i> strains (<i>cmu</i>B) with intraspecific indels that gave raise to four different alleles (<i>cmu</i>B1-4), which are exclusive of this species. Phylogenic analyses suggest that both proteins are present mainly in the <i>L. casei</i> and <i>Lactobacillus sakei</i> phylogenomic groups. A P40 ancestral gene was possibly present in the common ancestor of <i>Enterococcaceae</i>, <i>Lactobacillaceae</i> and <i>Streptococcaceae</i>. P75 is also present in <i>L. casei</i> and <i>L. sakei</i> groups, but its evolution is difficult to explain only by vertical transmission. Antibodies raised against the N-terminal regions of P40 and P75 improved their immunological detection in culture supernatants as they recognized almost exclusively proteins of <i>L. casei/paracasei/rhamnosus</i> strains, highlighting their structural similarity, that allowed to detect them in different fermented dairy products that contained probiotic <i>L. casei</i> strains. Purified P40 and P75 proteins showed no evident lytic activity but they complemented <i>L. casei</i> BL23 <i>cmu</i>A and <i>cmu</i>B defective mutants, respectively, thus proving that they actively participate in cell division.