Role of the Porphyromonas gingivalis iron-binding protein PG1777 in oxidative stress resistance.

Whole genome sequencing of the response of Porphyromonas gingivalis W83 to hydrogen peroxide revealed an upregulation of several uncharacterized, novel genes. Under conditions of prolonged oxidative stress in P. gingivalis, increased expression of a unique transcriptional unit carrying the grpE, dnaJ and three other hypothetical genes (PG1777, PG1778 and PG1779) was observed. The transcriptional start site of this operon appears to be located 91 bp upstream of the translational start, with a potential -10 region at -3 nt and a -35 region at -39 nt. Isogenic P. gingivalis mutants FLL273 (PG1777 : : ermF-ermAM) and FLL293 (PG1779 : : ermF-ermAM) showed increased sensitivity to and decreased survival after treatment with hydrogen peroxide. P. gingivalis FLL273 showed a fivefold increase in the formation of spontaneous mutants when compared with the parent strain after exposure to hydrogen peroxide. The recombinant PG1777 protein displayed iron-binding properties when incubated with FeSO4 and Fe(NH4)2(SO4).6H2O. The rPG1777 protein protected DNA from degradation when exposed to hydrogen peroxide in the presence of iron. Taken together, the data suggest that the grpE-dnaJ-PG1777-PG1778-PG1779 transcriptional unit may play an important role in oxidative stress resistance in P. gingivalis via its ability to protect against DNA damage.

Life in a Diverse Oral Community - Strategies for Oxidative Stress Survival.

BACKGROUND: While the oral cavity harbors more than 680 bacterial species, the interaction and association of selected bacterial species play a role in periodontal diseases. Bacterial species including Porphyromonas gingivalis, Treponema denticola and Tannerella forsythia, a consortium previously designated as the "red complex" is now being expanded to include other new emerging pathogens that are significantly associated with periodontal disease. HIGHLIGHT: In addition to novel mechanisms for oxidative resistance of individual species, community dynamics may lead to an overall strategy for survival in the inflammatory environment of the periodontal pocket. Complex systems controlled by response regulators protect against oxidative and nitrosative stress. CONCLUSION: The combination of these multifaceted strategies would provide a comprehensive defense and support system against the repetitive host immune response to promote microbial persistence and disease.

Protective role of the PG1036-PG1037-PG1038 operon in oxidative stress in Porphyromonas gingivalis W83.

As an anaerobe, Porphyromonas gingivalis is significantly affected by the harsh inflammatory environment of the periodontal pocket during initial colonization and active periodontal disease. We reported previously that the repair of oxidative stress-induced DNA damage involving 8-oxo-7,8-dihydroguanine (8-oxoG) may occur by an undescribed mechanism in P. gingivalis. DNA affinity fractionation identified PG1037, a conserved hypothetical protein, among other proteins, that were bound to the 8-oxoG lesion. PG1037 is part of the uvrA-PG1037-pcrA operon in P. gingivalis which is known to be upregulated under H2O2 induced stress. A PCR-based linear transformation method was used to inactivate the uvrA and pcrA genes by allelic exchange mutagenesis. Several attempts to inactivate PG1037 were unsuccessful. Similar to the wild-type when plated on Brucella blood agar, the uvrA and pcrA-defective mutants were black-pigmented and beta-hemolytic. These isogenic mutants also had reduced gingipain activities and were more sensitive to H2O2 and UV irradiation compared to the parent strain. Additionally, glycosylase assays revealed that 8-oxoG repair activities were similar in both wild-type and mutant P. gingivalis strains. Several proteins, some of which are known to have oxidoreducatse activity, were shown to interact with PG1037. The purified recombinant PG1037 protein could protect DNA from H2O2-induced damage. Collectively, these findings suggest that the uvrA-PG1037-pcrA operon may play an important role in hydrogen peroxide stress-induced resistance in P. gingivalis.

Oxidative stress resistance in Porphyromonas gingivalis.

Porphyromonas gingivalis, a black-pigmented, Gram-negative anaerobe, is an important etiologic agent of periodontal disease. The harsh inflammatory condition of the periodontal pocket implies that this organism has properties that will facilitate its ability to respond and adapt to oxidative stress. Because the stress response in the pathogen is a major determinant of its virulence, a comprehensive understanding of its oxidative stress resistance strategy is vital. We discuss multiple mechanisms and systems that clearly work in synergy to defend and protect P. gingivalis against oxidative damage caused by reactive oxygen species. The involvement of multiple hypothetical proteins and/or proteins of unknown function in this process may imply other unique mechanisms and potential therapeutic targets.

DNA repair of 8-oxo-7,8-dihydroguanine lesions in Porphyromonas gingivalis.

The persistence of Porphyromonas gingivalis in the inflammatory environment of the periodontal pocket requires an ability to overcome oxidative stress. DNA damage is a major consequence of oxidative stress. Unlike the case for other organisms, our previous report suggests a role for a non-base excision repair mechanism for the removal of 8-oxo-7,8-dihydroguanine (8-oxo-G) in P. gingivalis. Because the uvrB gene is known to be important in nucleotide excision repair, the role of this gene in the repair of oxidative stress-induced DNA damage was investigated. A 3.1-kb fragment containing the uvrB gene was PCR amplified from the chromosomal DNA of P. gingivalis W83. This gene was insertionally inactivated using the ermF-ermAM antibiotic cassette and used to create a uvrB-deficient mutant by allelic exchange. When plated on brucella blood agar, the mutant strain, designated P. gingivalis FLL144, was similar in black pigmentation and beta-hemolysis to the parent strain. In addition, P. gingivalis FLL144 demonstrated no significant difference in growth rate, proteolytic activity, or sensitivity to hydrogen peroxide from that of the parent strain. However, in contrast to the wild type, P. gingivalis FLL144 was significantly sensitive to UV irradiation. The enzymatic removal of 8-oxo-G from duplex DNA was unaffected by the inactivation of the uvrB gene. DNA affinity fractionation identified unique proteins that preferentially bound to the oligonucleotide fragment carrying the 8-oxo-G lesion. Collectively, these results suggest that the repair of oxidative stress-induced DNA damage involving 8-oxo-G may occur by a still undescribed mechanism in P. gingivalis.