Characterization of the Porphyromonas gingivalis Type IX Secretion Trans-envelope PorKLMNP Core Complex

Abstract

Edited by Thomas SöllnerThe transport of proteins at the cell surface of Bacteroidetes depends on a secretory apparatus known as type IX secretion system (T9SS). This machine is responsible for the cell surface exposition of various proteins, such as adhesins, required for gliding motility in Flavobacterium, S-layer components in Tannerella forsythia, and tooth tissue-degrading enzymes in the oral pathogen Porphyromonas gingivalis. Although a number of subunits of the T9SS have been identified, we lack details on the architecture of this secretion apparatus. Here we provide evidence that five of the genes encoding the core complex of the T9SS are co-transcribed and that the gene products are distributed in the cell envelope. Protein-protein interaction studies then revealed that these proteins oligomerize and interact through a dense network of contacts.

Quantitative proteomic analysis of the type IX secretion system mutants in Porphyromonas gingivalis

“…The next abundant proteins were PorG, PorP, PorE and Plug which all appeared to be of similar levels. PorK, PorL, PorM and PorN were ~10‐fold higher in abundance than Sov consistent with the known multimerization of these proteins (Gorasia et al, ; Vincent et al, ) (Figure ). PorU, PorQ and PorZ were of similar abundance to PorK, PorL, PorM and PorN.…”

Type IX secretion: the generation of bacterial cell surface coatings involved in virulence, gliding motility and the degradation of complex biopolymers

“…Topological predictions suggest that PorL is predominantly cytoplasmic while PorM is suggested to be predominantly periplasmic (Shrivastava et al ., ). The cytoplasmic localisation of the soluble PorL domain was confirmed by its binding to the cytoplasmic response regulator, PorX (Vincent et al ., ). Furthermore, cysteine accessibility assays confirmed the localizations of PorL and PorM as depicted in Fig.…”

“…Furthermore, cysteine accessibility assays confirmed the localizations of PorL and PorM as depicted in Fig. (Vincent et al ., ). PorL and PorM have been shown to bind to each other (Gorasia et al ., ; Vincent et al ., ) and recently, PorM was shown to also interact with PorK and PorN (Vincent et al ., ).…”

“…Besides the putative regulatory components (see below), there are seven further proteins with experimental evidence for involvement in the T9SS. In P. gingivalis , PorP has been modelled as a 14‐stranded OM β‐barrel similar to PorV and PorQ and was found associated with PorK and PorM in E. coli by immunoprecipitation experiments (Vincent et al ., ). PorP however was not required for the formation of large complexes comprising PorK, PorL, PorM and PorN (Sato et al ., ) and moreover, the abundance of PorP in wild type cells is approximately 20‐fold lower than these other four components (unpublished data).…”

“…In P. gingivalis , the porPKLMN genes were shown to be co‐transcribed (Vincent et al ., ) however, quantitative proteomics indicates that PorP is more than an order of magnitude less abundant than PorK, PorL, PorM and PorN raising the possibility that there is an additional promoter to drive transcription of the porKLMN unit alone (unpublished data). PorK was predicted to be a lipoprotein and was localised to the OM along with PorN by differential detergent solubility of the IM and OM, while PorL and PorM were localised to the IM (Sato et al ., ; Shrivastava et al ., ; Vincent et al ., ). Blue Native gel electrophoresis (BN‐PAGE) and Western blotting revealed that all four of these proteins formed large complexes of over 1 MDa consistent with them forming a secretion apparatus spanning both membranes (Sato et al ., ).…”