Hemin binding by Porphyromonas gingivalis strains is dependent on the presence of A‐LPS

Abstract

Porphyromonas gingivalis is a Gram‐negative black pigmenting anaerobe that is unable to synthesize heme [Fe(II)‐protoporphyrin IX] or hemin [Fe(III)‐protoporphyrin IX‐Cl], which are important growth/virulence factors, and must therefore derive them from the host. Porphyromonas gingivalis expresses several proteinaceous hemin‐binding sites, which are important in the binding/transport of heme/hemin from the host. It also synthesizes several virulence factors, namely cysteine‐proteases Arg‐ and Lys‐gingipains and two lipopolysaccharides (LPS), O‐LPS and A‐LPS. The gingipains are required for the production of the black pigment, μ‐oxo‐bisheme {[Fe(III)PPIX]2 O}, which is derived from hemoglobin and deposited on the bacterial cell‐surface leading to the characteristic black colonies when grown on blood agar. In this study we investigated the role of LPS in the deposition of μ‐oxo‐bisheme on the cell‐surface. A P. gingivalis mutant defective in the biosynthesis of Arg‐gingipains, namely rgpA/rgpB, produces brown colonies on blood agar and mutants defective in Lys‐gingipain (kgp) and LPS biosynthesis namely porR, waaL, wzy, and pg0129 (α‐1, 3‐mannosyltransferase) produce non‐pigmented colonies. However, only those mutants lacking A‐LPS showed reduced hemin‐binding when cells in suspension were incubated with hemin. Using native, de‐O‐phosphorylated and de‐lipidated LPS from P. gingivalis W50 and porR strains, we demonstrated that hemin‐binding to O‐polysaccharide (PS) and to the lipid A moiety of LPS was reduced compared with hemin‐binding to A‐PS. We conclude that A‐LPS in the outer‐membrane of P. gingivalis serves as a scaffold/anchor for the retention of μ‐oxo‐bisheme on the cell surface and pigmentation is dependent on the presence of A‐LPS.

Using Tn-seq To Identify Pigmentation-Related Genes of Porphyromonas gingivalis: Characterization of the Role of a Putative Glycosyltransferase

“…The altered LPS structure, most notably the lack of APS, also likely contributes to the decreased heme binding and lack of pigmentation observed in the PG_0264 mutant. In fact, APS of P. gingivalis was recently shown to be critical in the binding of -oxo-bisheme and is required for pigmentation (31). In addition, it is possible that PGN_0361/PG_0264 directly glycosylates Rgp, which affects its catalytic activity (32).…”

Isolates fromMonechma ciliatumseeds' extract hamperedPorphyromonas gingivalishemagglutinins

“…P. gingivalis TDC60 (Watanabe et al, ) was provided by the RIKEN BioResource Center through the National Bio‐Resource Project of the Ministry of Education, Culture, Sports, Science and Technology, Japan. The bacterium was grown in Tryptic Soy Blood Agar medium (Agar Base EH) supplemented with hemin (5 mg/L), menadione (1 mg/L), and horse blood at 37°C under anaerobic conditions for 4–5 days (Gao et al, ; Rangarajan, Aduse‐Opoku, Paramonov, Hashim, & Curtis, ). The cells were then inoculated into 5 ml of GAM broth medium and incubated under anaerobic conditions at 37°C for 2–3 days until the middle stationary phase of bacterial growth.…”

Importance of heterogeneity in Porhyromonas gingivalis lipopolysaccharide lipid A in tissue specific inflammatory signalling

“…P. gingivalis cannot synthesize the protoporhyrin IX ring and does not have siderophores for alternative bioavailability of hemin [36–39]. Therefore, P. gingivalis depends on the host for a supply of heme [40]. The lysis of multiple erythrocytes, in order to feed P. gingivalis heme, would lead to decrease in oxygen for the host, which would in turn contribute to ischemia.…”

“…µ-Oxo-bisheme pigmentation is a form of a bacteriocin and a virulence factor of P. gingivalis [40]. Both Arg and Lys gingipains are required for the deposition of this characteristic black pigment.…”

“…Both Arg and Lys gingipains are required for the deposition of this characteristic black pigment. Absence of A-LPS in the extracellular surface of P. gingivalis diminishes the scaffold/anchoring mechanism that otherwise retains Arg- and Lys-gingipains [40]. This implies that A-LPS serves as a site for deposition/binding of hemin.…”