Role of gingipains in growth of Porphyromonas gingivalis in the presence of human serum albumin

Citation
D. Grenier et al., Role of gingipains in growth of Porphyromonas gingivalis in the presence of human serum albumin, INFEC IMMUN, 69(8), 2001, pp. 5166-5172
Citations number
26
Categorie Soggetti
Immunology
Journal title
INFECTION AND IMMUNITY
ISSN journal
00199567 → ACNP
Volume
69
Issue
8
Year of publication
2001
Pages
5166 - 5172
Database
ISI
SICI code
0019-9567(200108)69:8<5166:ROGIGO>2.0.ZU;2-1
Abstract
Porphyromonas gingivalis, a bacterium associated with active chronic period ontitis lesions, produces several proteolytic enzymes that are thought to b e involved in host colonization, perturbation of the immune system, and tis sue destruction. The aim of the present study was to investigate the contri bution of Arg- and Lys-gingipains produced by P. gingivalis to its growth. Although all of the proteins studied were degraded by P. gingivalis, only h uman serum albumin and transferrin supported growth during serial transfers in a chemically defined medium (CDM). Growth studies with site-directed gi ngipain-deficient mutants of P. gingivalis revealed that inactivation of bo th gingipains prevents growth, whereas inactivation of either Arg- or Lys-g ingipain activity extended the doubling times to 33 or 13 h, respectively, compared to 9 h for the parent strain. Growth of the mutants and the parent strain was similar when the CDM was supplemented with a protein hydrolysat e instead of human serum albumin. Incubation of resting P. gingivalis ATCC 33277 cells with fluorophore-labeled albumin indicated that the proteolytic fragments generated by the gingipains were internalized by the bacterial c ells. Internalization of fluorophore-labeled albumin fragments was reduced or completely inhibited in the proteinase-deficient mutants. Interestingly, gingival crevicular fluid samples from diseased periodontal sites containe d low-molecular-mass albumin fragments, whereas samples from healthy sites did not. The critical role of proteinases in the growth of P. gingivalis wa s further investigated using specific Arg- and Lys-gingipain inhibitors. Ad ding the inhibitors to CDM containing albumin revealed that leupeptin (Arg- gingipain A and B inhibitor) was more efficient at inhibiting growth than c athepsin B inhibitor II (Lys-gingipain inhibitor). Our study suggests that Arg-gingipains and, to a lesser extent, Lys-gingipain play an important rol e in the growth of P. gingivalis in a defined medium containing a human pro tein as the sole carbon and nitrogen source.