Novel protein identification methods for biomarker discovery via a proteomic analysis of periodontally healthy and diseased gingival crevicular fluid samples

Aim: To identify possible novel biomarkers in gingival crevicular fluid (GCF) samples from chronic periodontitis (CP) and periodontally healthy individuals using high-throughput proteomic analysis. Materials and Methods: Gingival crevicular fluid samples were collected from 12 CP and 12 periodontally healthy subjects. Samples were trypically digested with trypsin, eluted using high-performance liquid chromatography, and fragmented using tandem mass spectrometry (MS/MS). MS/MS spectra were analysed using PILOT_PROTEIN to identify all unmodified proteins within the samples. Results: Using the database derived from Homo sapiens taxonomy and all bacterial taxonomies, 432 human (120 new) and 30 bacterial proteins were identified. The human proteins, angiotensinogen, clusterin and thymidine phosphorylase were identified as biomarker candidates based on their high-scoring only in samples from periodontal health. Similarly, neutrophil defensin-1, carbonic anhydrase-1 and elongation factor-1 gamma were associated with CP. Candidate bacterial biomarkers include 33 kDa chaperonin, iron uptake protein A2 and phosphoenolpyruvate carboxylase (health-associated) and ribulose biphosphate carboxylase, a probable succinyl-CoA:3-ketoacid-coenzyme A transferase, or DNA-directed RNA polymerase subunit beta (CP-associated). Most of these human and bacterial proteins have not been previously evaluated as biomarkers of periodontal conditions and require further investigation. Conclusions: The proposed methods for large-scale comprehensive proteomic analysis may lead to the identification of novel biomarkers of periodontal health or disease.