Mining the oncoproteome and studying molecular interactions for biomarker development by 2DE, ChIP and SPR...

This article provides a comprehensive updated review of three proteomics technologies (2D gel electrophoresis [2DE]; chromatin immunoprecipitation [ChIP] and its related alternates, and surface plasmon resonance [SPR]), and their use in cancer biomarker discovery and studying molecular interactions. 2DE proteomics has an advantage in visualizing changes in the intact molecular weight and isoelectric point (pI) of a protein, which reflect biologically significant processing and charge alterations in addition to post-translation modifications. However, proteins that are hydrophobic, low abundance or with extreme pIs or molecular weight, are poorly represented. Despite these shortfalls, improvements have been made that provide enhanced resolution by use of immobilized pH gradients (IPGs), narrow strip large 2DE gels, and serial and parallel formats. Prefractionation of the sample is essential for studying low-abundance markers, although it subjects the experimental design to additional nonbiologic variations. 2DE has been applied to the successful identification of oncoproteins in serum and tissues of cancers, such as colon, breast, prostate and pancreas. Interactions between protein and DNA are essential for cellular function. The process of developing global approaches to studying chromatin began with the in vitro characterization of chromatin structural components by the versatile ChIP assay and its modifications, which are capable of analyzing protein-DNA interactions in vivo. These methods have been used to assess gene expression, transcription factor binding and/or histone modifications in various organisms, including human. Label-free, real-time analysis of interactions in biological systems employing SPR biosensors that measure changes in refractive index of solvents near the surface during complex formation or dissociation of proteins, oligonucleotides, oligosaccharides and lipids to small molecules, phage, viral particles and cells is used extensively in research laboratories and the pharmaceutical and biotechnology industries for studying drug discovery, biopharmaceutical developments and clinical immunogenicity assays, as well as in manufacturing and quality-control methods.