Proteomic analyses of the two mucus layers of the colon barrier reveal that their main component, the Muc2 mucin, is strongly bound to the Fcgbp protein

The colon epithelium is protected from the luminal microbes as recently revealed by an inner firmly attached mucus layer impervious to bacteria and an outer loose mucus layer that is the habitat of bacteria. For an additional understanding of these layers, we analyzed the protein composition of these two mucus layers from the mouse colon. Proteomics using nano-LC-MS and MS/MS revealed more than 1000 protein entries. As the mucus layers contain detached cells, a majority of the proteins had an intracellular origin. However, at least 44 entries were described as secreted proteins and predicted to be mucus constituents together with extracellular/plasma and Bacterial proteins, the latter largely in the loose mucus layer. A major protein was the Muc2 Mucin that by its net-like disulfide-bonded polymer structure builds the mucus. When guanidinium chloride insoluble Muc2 units were analyzed, N-terminal parts of the Fc-gamma binding protein (Fcgbp) was found to be covalently attached in mouse and human colon, whereas its C-terminus was lost by reducing the disulfide bonds. In conclusion, the Fcgbp protein is probably cleaved at GD/PH and covalently attached to Muc2 via one or several of its von Willebrand D domains.