Human_MCF7_membrane - Searching Missing Proteins Based on the Optimization of Membrane Protein Enrichment and Digestion Process

MCF7 cell was an important stabilized breast cancer cell in cancer research. A membrane proteomic method was developed and compared in this study. A membrane protein enrichment method composed of ultra-centrifugation and detergent-based extraction was first developed. Then in-solution digestion with detergents and eFASP (enhanced filter aided sample preparation) with detergents were compared with the time consuming in-gel digestion method. Our data indicated that ultra-centrifugation combined detergent-based extraction can effectively enrich membrane proteins with higher purity. Among the in-solution digestion methods, the eFASP combined with RapiGest method identified 1125 membrane proteins. Similarly, the eFASP combined with SDC identified 1069 membrane proteins, however, the in-gel digestion characterized 1091 membrane proteins. Totally, with the 5 digestion methods, 1390 membrane proteins were identified with ≥1 unique peptides, among which 1345 membrane proteins contain unique peptides ≥2. This is the biggest membrane protein datasets for MCF7 cell line and even breast cancer tissue samples. Interestingly, we realized that 9 missing proteins (MPs) present with one or more unique peptides. After verification, 4 of them were validated by synthesized peptide with amino acids larger than 9.