Identification of a novel Porphyromonas gingivalis outer membrane protein, PG534, required for the production of active gingipains

Gingipains are secreted endopeptidases important for the virulence and proliferation of Porphyromonas gingivalis; however, their secretion and biogenesis process is not yet fully elucidated. The PG0534 gene of P. gingivalis W83 encodes a novel protein, PG534, of unknown function. In a PG0534 deletion mutant 83K25, the activities of Arg-gingipains (RgpA and RgpB) and Lys-gingipain (Kgp) were reduced to 4-22% of those of the wild-type W83, while the activities of secreted exopeptidases DPPIV, DPP-7, and PTP-A were unaffected. This indicates that PG534 is required for the gingipain activity. Immunoblot analysis using anti-Rgp or anti-Kgp antiserum showed that abnormal forms of gingipains were detected in the extracellular fraction from 83K25, suggesting that 83K25 exhibits dysfunctional gingipain secretory activity. Normal carbohydrate biogenesis of lipopolysaccharide is required for production of the active gingipains; however, lipopolysaccharide was not deficient in 83K25. Subcellular fractionation and immunoblot analysis using anti-PG534 antiserum localized PG534 to the outer membrane. In conclusion, we identified PG534 as a novel outer membrane protein required for the biogenesis of gingipains.