The gene encoding PGN_1476 in the PorSS-deficient strain was expressed about three times more than that in the PorSS- proficient strain. As the relative amounts of the protein spots were < 20% (Table 2), the results suggest that decrease of the 10 secreted proteins in the PorSS-deficient mutant are

mostly dependent on the defect in the PorSS. The 10 PorSS-dependently secreted proteins as well as precursor Tacrolimus mw forms of Arg-gingipains (RgpA and RgpB) and Lys-gingipain (Kgp) had CTDs in which the conserved DxxG and GxY motifs and the conserved Lys residue are located (Seers et al., 2006; Fig. 5). Seers et al. (2006) reported that 34 CTD family proteins with sequence similarity to the C-terminal region of the RgpB precursor

were identified by a blast search with the P. gingivalis W83 genome, which include the 10 proteins identified in the present Obeticholic Acid in vitro study. Slakeski et al. (2010) suggested that the CTD of RgpB is essential for covalent attachment to the cell surface by an A-LPS anchor containing anionic polysaccharide repeating units. In our previous studies (Kondo et al., 2010; Shoji et al., 2011), we demonstrated that HBP35 and TapA were modified by A-LPS and anchored on the bacterial cell surface. In addition, the green fluorescent protein–CTD fusion study revealed that the CTDs of CPG70, PAD and HBP35 as well as RgpB play roles in PorSS-dependent translocation and glycosylation (Shoji et al., 2011). We suggested in the study both that the CTD region functions as a recognition signal for the PorSS and that glycosylation of CTD proteins occurs after removal of the CTD region. Cleaved CTD fragments of HBP35, CPG70, PAD, RgpB and PGN_1767 have recently been found in the culture supernatants of P. gingivalis (Glew et al., 2012), which is consistent with the present study and supports

our model (Shoji et al., 2011). Our results strongly indicate that the P. gingivalis secreted proteins with CTDs, which are responsible for colony pigmentation, hemagglutination, adherence and modification/processing of the bacterial surface proteins and host Olopatadine proteins, are translocated to the cell surface by the PorSS. In the present study, using 2D-PAGE and MS we identified 10 proteins secreted into the extracellular milieu by the PorSS. All of the proteins possessed CTDs. They included HBP35 in heme acquisition, TapA in virulence, PAD in citrullination of C-terminal Arg residues of the surface proteins and CPG70 in processing of C-terminal Arg and Lys residues. These results indicate that the PorSS is used for secretion of a number of proteins other than gingipains and that the CTDs of the proteins are associated with the PorSS-dependent secretion.