The Study Of Several Methods On Proteomics

Recent developments in analytical methods for protein characterization, and the growing rate at which whole genome sequencing projects are being completed, have combined to support the emergence and development of the new field of 'proteomics' . Proteomics, the study of protein expression and function on a genome scale, is the amalgamation of very many different experimental approaches ranging from the analysis of protein interactions by genome-wide two hybrid screening, to more direct analysis of protein expression, sequence and structure. Proteome indicates the PROTEins expressed by a genome. The technology of proteomics requires the separation, identification of large number of proteins. In proteome projects, which aim to identify and characterize all proteins expressed by an organism or tissue, the identidfication of proteins is critical. The related techniques and methods were developing quickly. But considering the complexity of proteomics and the limit of current techniques, there are still lots of problems needed to be resolved. Three methods was studied in this paper, some results are as follows. 1. MADLI-TOF-PSD MethodDue to its very short analysis time, its high sensitivity and easy of automation, MALDI-peptide mass fingerprinting (PMF) has become the preferred method for identifying proteins of which the sequences are available in database. Unfortunately, PMF can also lead to ambiguous protein identification because of insufficient protein sequence coverage, complex mixtures of tryptic peptides from co-migrating proteins and novel protein of no match in database. To avoid this problem, post-sourcedecay (PSD) MALDI mass spectrometry was developed for high-sensitivity peptide sequencing. Moreover, two N~terminal chemical modified reagent was studied .This modification significantly facilitates sequence interpretation by providing strong intensity C-terminal product ions(y-type ions) in the matrix-assisted laser desorption/ionization -postsource decay (MALDI-PSD)-MS. We can easy to read the sequence information by distinct difference between the adjacent y-type ion peaks.2. The application of surface plasmon resonance technology in proteomics studyBiomolecular interaction analysis mass spectrometry (BIA/MS) is an approach combining the surface plasmon resonance(SPR) technology and mass spectrometry for the real-time analysis and identification of interaction molecules that interact specifically with a known immobilized ligand. BIA/MS is a very promising method applied in proteomics for the characterization of protein-protein interactions.. In this study, by using the Biacore-X instrument, the method was successfully applied to the model protein microglobulin interacting with its antibody, a unambiguous identification was obtained at fmol level. In addition, we studied the function of some toxin by using the SPR technology.3. The method of quantitative via Isotope-code affinity tagIn order to reach the goal of quantitative proteomics, isotope-coded affinity tags(ICATs) were employed to identify and quantitate changes in protein expression. Firstly, The method was applied to analyze the standard peptide and protein to optimize the procedure of ICAT. After the protein was labled, trypsin was applied, two-dimensional liquidchromatography-mass spectrometry coupled with strategy was used in analysis . The result revealed that this strategy facilitated the fast and accurate quantitation by one-step and online desalting, isolation and identification of labeled peptides.