Abstract
Advances in Proteome Analysis by Mass Spectrometry
Highlights
This review describes recent, innovative advances in mass spectrometry-based proteome analysis that collectively have converged into a generic new approach to proteomics
Tive technique and, without the availability of reliable tools for the identification of the separated protein species, of limited utility as a molecular biology research tool. This changed in the early 1990s when two revolutionary techniques, matrix-assisted laser desorption ionization (MALDI) timeof-flight (TOF) mass spectrometry (MS) (13, 14) and electrospray ionization (ESI) (15, 16) MS and tandem mass spectrometry (MS/ MS) replaced the slower and less sensitive chemical degradation methods (17, 18) as the methods of choice for the identification of proteins separated by 2DE (19 –24)
Protein identification was accomplished using peptide-mass fingerprinting by MALDI-TOF MS, as initially described by Henzel et al (26) and independently by others, nano-ESI tandem mass spectrometry (MS/MS) (27, 28), or reverse-phase (RP) microcapillary liquid chromatography (LC) ESI MS/MS (29 –34) using automated, data-dependent scanning and dynamic exclusion of peptide ions already analyzed in the same experiment (29, 35– 37)
Summary
The interpretation of the information contained in the genomic sequence of a species with respect to the structure, function, and control of biological processes is a technical and conceptual challenge for current research methods. The three components permit the relative measurement of abundance and the identification of the components of very complex protein mixtures rapidly and with a high degree of automation, without the need to separate protein mixtures prior to analysis Variations of this technology have the potential to systematically and quantitatively determine properties of proteins that reflect their functional state. The challenge facing any comprehensive proteomics approach is one of separating and simplifying very complex mixtures of proteins in which individual components differ in abundance by six or more orders of magnitude, while retaining enough information to allow for comprehensive characterization of expressed proteins To this end, the combination of selective labeling of proteins with stable isotope-containing affinity reagents and multidimensional liquid chromatography in conjunction with automated, data-dependent tandem mass spectrometry and sequence data base searching has proven effective and has been shown to overcome at least some of the critical limitations of the 2DE-MS-based approach to proteomics
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