Immunoproteomic analyses of outer membrane proteins of Mannheimia haemolytica and identification of potential vaccine candidates

Immunoproteomic analyses were used to characterize the outer membrane proteome of Mannheimia haemolytica, formerly Pasteurella haemolytica, serotype 1, and determine potential vaccine candidate proteins. 2-DE of M. haemolytica outer membranes was followed by immunoblot analyses using naive and convalescent bovine sera. Proteins were identified using MALDI-TOF and LC-MS/MS. Spectral data was used to mine M. haemolytica protein database and 132 immunoreactive proteins were identified. Bioinformatic analysis using PSORTb, SubLoc, LipoP, BOMP, MCMBB, and TMB-Hunt/BBTM to predict subcellular localization of immunoreactive proteins and beta-barrels narrowed the list down to 55 candidates. Functional characterization of 55 proteins predicted 16 (29%) are involved in cell structure, 13 (23.6%) in transport/virulence, ten (18.2%) as unknown, six (10.9%) in general metabolism, four (7.27%) in cell process, two (3.64%) in translation, and one (1.8%) each in DNA replication, regulation, transcription, and virulence. Prediction of beta-barrel formation was between 11 and 31 immunoreactive proteins depending on the bioinformatic tool employed. Some of these proteins have potentials to be developed into stand-alone vaccines or components of vaccines. Of those proteins, several have already been characterized. Finally, although characteristics of many of M. haemolytica immunoreactive proteins identified in this study were obtained from published data and predictions using bioinformatics tools, five proteins previously listed in the published M. haemolytica sequence as unidentified were found to have correlates with functional proteins in other bacterial species.