Enrichment of Cell Surface-Associated Proteins in Gram-Positive Bacteria by Biotinylation or Trypsin Shaving for Mass Spectrometry Analysis

In microbial cells surface-exposed proteins represent a physiologically important class of molecules as they enable these cells to interact with their environment both as free-living organisms and during interactions with a host. However, the characteristics of these proteins are quite divergent, which makes attempts to enrich, analyze, and quantify these molecules a challenging task. In this chapter two complementary methods for the enrichment and identification of cell surface-associated proteins, namely the biotinylation and the shaving approaches, are presented. Both protocols have been optimized for Gram-positive bacteria, and we provide a step-by-step guide for sample generation. Possible pitfalls during protein preparation are discussed.