Porphyromonas gingivalis is a nonmotile pathogen associated with chronic periodontitis. P. gingivalis secretes virulence proteins such as the Arg- and Lys-gingipain proteinases via a poorly characterized system that is dependent on substrates having a carboxyl-terminal motif, the CTD. The secretion system has paralogues in other members of the Bacteroidetes phylum including Flavobacterium johnsoniae where it is needed for motility. P. gingivalis secreted proteins are modified with anionic lipopolysaccharide (A-LPS) which is thought to be coordinately attached during secretion. We identified the 75 kDa protein PG1058 as a potential P. gingivalis secretion system component. Objectives: This study examined characteristics of PG1058 and its function in P. gingivalis.
Methods: We deleted pg1058 of P. gingivalis and complemented the mutant by allele replacement. We examined growth characteristics of both strains and the distribution of A-LPS and CTD-substrate proteins using western immunoassays and enzyme assays. Molecular modeling was used to predict the PG1058structure.
Results: The pg1058 mutant grew slowly, was unable to pigment on blood agar and had no Arg-X or Lys-X amidolytic activity thus indicating no gingipain protease function. Westerns showed the CTD-substrate proteins were accumulating in the periplasm and they were not A-LPS-modified; A-LPS was atypically abundant in the pg1058 mutant culture supernatant. Complementation of the pg1058 mutation restored the phenotype to wild-type. Modelling PG1058 revealed a novel architecture comprising 4 domains, (I) a tetratricopeptide repeat (TPR, a protein-protein interaction motif), (II) a beta-propeller TolB-like domain (typically involved in substrate-binding), (III) MotY-N which forms part of Vibrio flagellar rotor and (IV) a domain resembling peptidoglycan-binding domains including MotY-C. PG1058 may anchor to the P. gingivalis peptidoglycan via MotY domains and interact with proteins via the TPR and TolB-like domains.
Conclusion: PG1058 is needed for P. gingivalis CTD-protein secretion and modification supporting that it is part of the P. gingivalis protein secretion system.