Label-Free Quantitative Proteomics of Oral Microbial Communities

The oral cavity is a habitat for different microorganisms, of which bacteria are best described. Studying different bacterial taxa and their proteins is crucial to understanding their interactions with the host and other microbes. Also, for bacteria with virulence potential, identifying novel antigenic proteins is essential to finding candidates for the development of vaccines. Here, a workflow for gel-free and label-free protein analysis of oral bacterial species grown in vitro as a biofilm and a planktonic culture is described. Details on cultivation, protein extraction and digestion, peptide cleanup, LC-MS/MS run parameters, and subsequent bioinformatics analysis are included. Challenging steps in the workflow, such as growing different types of bacteria and selecting a suitable protein database, are also discussed. This protocol provides a valuable guide for metaproteomic experiments using multi-species models of oral bacteria.