Escherichia coli is a diverse species of bacteria. Some isolates of E. coli can cause hemorrhagic uremic syndrome in humans through consumption of contaminated food. We hypothesize that secreted and surface-associated proteins in E. coli may contribute to bacterial persistence in food products and to pathogenicity in humans. Here we report the proteomic analyses of secretomes from two serotypes of E. coli, O157:H7 and O104:H4, that cause human foodborne illness. The secreted proteins were isolated from the liquid cultures and analyzed by liquid chromatography-tandem mass spectrometry. We report 57 secreted proteins for O157:H7 and 67 for O104:H4 with increased abundance in minimal medium. As signature proteins, O157:H7 expressed intimin and translocated intimin receptor. By contrast, O104:H4 expressed enteroaggregative fimbriae, serine protease autotransporters, Shiga toxin, and beta-lactamase. The E. coli O104:H4 released more cytosolic proteins into the culture media than the O157:H7 serotype, indicating that it might be a leaky strain. The differential abundance of two transpeptidases YbiS and YnhG, along with other lipoproteins in O157:H7 indicates that this serotype secretes several peptidol-lipoprotein components of biofilm. In addition, we observed differential abundance of several ABC transporters, proteases and lipoproteins, which give each strain a unique survival strategy and drug resistance. The identification of secreted proteins will enable us understand better how these bacteria persist in food products and cause human disease.