Porphyromonas gingivalis C-terminal signal peptidase PG0026 and HagA interact with outer membrane protein PG27/LptO.

Outer membrane protein PG27 is essential for secretion/maturation of conserved C-terminal domain (CTD) proteins such as gingipains, HagA, and PG0026. To determine the binding partner(s) of PG27, we used a Porphyromonas gingivalis mutant strain, 83K48, which expressed functional histidine-tagged PG27. Purification of histidine-tagged PG27 from 83K48 found that 136-kDa and 264-kDa proteins accompanied histidine-tagged PG27. Mass spectrometry revealed the 136-kDa protein and 264-kDa protein to be PG0026 and PG1837 (HagA), respectively. PG0026 is a C-terminal signal peptidase which cleaves the CTDs of other CTD proteins. A cross-linking and a native electrophoresis studies suggested the interaction between histidine-tagged PG27 and HagA and the interaction between histidine-tagged PG27 and PG0026. We constructed Porphyromonas gingivalis gene disrupting mutants, and characterized them. PG0026 was required for the full activities of gingipains, whereas HagA was not. A mutation disrupting PG0026 affected localization of PG27, but a mutation disrupting PG1837 showed little effect on the expression and localizations of PG27 and PG0026. To determine the functional role of HagA, we used PG1837-disruptant 83K54 which expressed functional histidine-tagged PG27. Histidine-tagged PG27 in 83K54 was co-purified with not only PG0026 but also several contaminated proteins. These results suggest that PG0026 interacts with PG27 in the absence of HagA, and that the binding state of a PG27-PG0026 complex was affected and stabilized by HagA. Taken together, these data suggest that secreted PG0026 anchors to the cell by interacting with PG27, is stabilized by HagA, and functions in processing of other CTD proteins such as gingipains.