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Purification and characterization of fibroblast-activating factor isolated from Porphyromonas gingivalis W50
1Department of Periodontics, University of Texas Health Science Center, San Antonio 78284-7894.
Abstract:
A 24-kDa polypeptide which activated the incorporation of [3H]thymidine into human fibroblasts was isolated from the outer membrane vesicles of Porphyromonas gingivalis W50. This polypeptide, named fibroblast activating factor (FAF), was isolated by 3-[(3-cholamidopropyl)-dimethyl-ammonio]-1-propane-sulfonate (CHAPS) detergent extraction and purified by DEAE ion-exchange chromatography and preparative isoelectric focusing. Purified FAF (100 ng of protein per ml) caused a 400% increase in [3H]thymidine incorporation into human gingival fibroblasts (HGFs) compared with results for controls. FAF was characterized as (i) a polypeptide with molecular masses of 24 kDa when heated at 100 degrees C for 5 min and 44 kDa when unheated, (ii) heat sensitive but not affected by selected reducing reagents, and (iii) possessing slight phosphatase activity. N'-terminal sequence analysis revealed no homology with P. gingivalis peptides or with any host-derived growth factors. These data suggest that FAF functions as a significant virulence factor which in vivo is capable of modulating homeostasis in local connective tissues.
Insights
A novel polypeptide, fibroblast activating factor (FAF), was isolated from Porphyromonas gingivalis. This potent virulence factor significantly increases human fibroblast proliferation, suggesting a role in tissue modulation.
Area of Science:
- Microbiology
- Molecular Biology
- Biochemistry
Background:
- Porphyromonas gingivalis is a key pathogen in periodontal disease.
- Bacterial outer membrane components can influence host cell behavior.
Purpose of the Study:
- To isolate and characterize a polypeptide from P. gingivalis that affects human fibroblast activity.
- To investigate the role of this factor as a potential virulence determinant.
Main Methods:
- Isolation of a 24-kDa polypeptide (FAF) from P. gingivalis outer membrane vesicles.
- Purification using CHAPS detergent extraction, ion-exchange chromatography, and isoelectric focusing.
- Assessment of FAF's effect on [3H]thymidine incorporation in human gingival fibroblasts (HGFs).
Main Results:
- Purified FAF significantly increased [3H]thymidine incorporation in HGFs by 400%.
- FAF is a heat-sensitive polypeptide with molecular masses of 24 kDa (heated) and 44 kDa (unheated).
- N'-terminal sequencing showed no homology to known bacterial or host factors.
Conclusions:
- FAF is a novel P. gingivalis virulence factor that stimulates fibroblast proliferation.
- This activity suggests FAF modulates local connective tissue homeostasis during infection.
- FAF represents a potential target for understanding and treating P. gingivalis-associated pathologies.