Identification of outer membrane proteins of Porphyromonas gingivalis 47A-1 by Shotgun proteomics method

Objective: To identify outer membrane proteins of Porphyromonas gingivalis (P. g.) by shotgun proteomics method so as to lay a foundation for further study of pathogenicity and proteomics of P. g. Methods: P. g47A-1 were cultured and collected when they were in good conditions. Outer membrane proteins of Pg 47A-1 were extracted by differential centrifugation and separated by SDS-PAGE. We selected trypsin for protein hydrolysis in SDS-PAGE gel, separated the peptide fragments using cap HPLC, and analyzed each peptide fragment in MS/MS. With the help of Sequest software, the NCBI database was searched for the identification of outer membrane proteins of P.g47A-1. Results: The proteins in gels were stained using Coomassie Brilliant Blue, and the outer membrane protein bands were mainly located between 14.4 and 97 ku. Separation of the peptides was achieved by capillary high performance liquid chromatography and identified using the total ion flow diagram. Peptide fragment sequence showed that a total of 623 peptides, 136 differential proteins were identified, of which 51 proteins were known and the other 85 proteins were unknown. Conclusion: Shotgun technique can analyze a large number of unknown proteins in P.g47A-1, especially outer membrane proteins with low abundance and strong hydrophobicity. Compared with two-dimensional gel electrophoresis, "shotgun proteomics" might be a better method for the analysis of outer membrane proteins. The findings on OMPS of P. g in the present study might supply some clues for further understanding of its pathogenic mechanisms.