Proteomics protein structure analysis

The focus in proteomics is still on protein discovery and protein structure analysis rather than on dynamic proteomics and protein function. 

Jolles, Pierre, and Hans Jornvall. Proteomics in Functional Genomics Protein Structure Analysis. EXS, no. 88. Boston Birkhauser Verlag, 2000. 

A new chapter on the primary structure of proteins, which provides coverage of both classic and newly emerging proteomic and genomic methods for identifying proteins. A new section on the appHcation of mass spectrometry to the analysis of protein structure has been added, including comments on the identification of covalent modifications. 

ExPASy Proteomics tools (http //expasy.org/tools/), tools and online programs for protein identification and characterization, similarity searches, pattern and profile searches, posttranslational modification prediction, topology prediction, primary structure analysis, or secondary and tertiary structure prediction. 

The marriage of HPLC to mass spectrometry (MS), now developed into a mature instrumentation, continues to greatly impact many of the separation sciences, especially in pharmaceutical analysis where it has been used in new drug discovery [23,24] and in drug metabolite identification [25-27]. HPLC-MS has also made an impact on lipid research, providing a convenient approach to the analysis of phospholipids and fatty acids [28,29]. It has also greatly benefited the field of proteomics [30-34], especially analysis of protein structure and function. 

A variety of methods are available to detect proteins separated by electrophoresis or to measure the concentration of total protein in a solution. These methods are normally based on the binding of a dye to one of the amino acids in protein, or a color reaction with an amino acid side chain. The most commonly used stains for protein detection on gels are Coomassie Brilliant Blue (98) and silver stain (99,100). These methods detect any protein residues, either in solution or on an electrophoresis gel. Their main requirement is sensitivity, not specificity. New, more sensitive dyes are being developed for the proteomic analysis of protein structure and sequence, for example Ruby Red (101). 

General consensus has sub-divided proteomics into three main areas, Expression Proteomics, Functional Proteomics, and Structural Proteomics. Expression Proteomics (sometimes called differential-expression proteomics) involves the analysis of differential protein expression by protein... 

Proteome refers to protein complement expressed by a genome. Thus proteomics concerns with the analysis of complete complements of proteins. It is the study of proteins that are encoded by the genes of a cell or an organism. Such study includes determination of protein expression, identification and quantification of proteins as well as characterization of protein structures, functions and interactions. The functional classification of proteins in genomes (i.e., proteomes) can be accessed from the Proteome Analysis Database at http //ebi.ac.uk/proteome/ (Apweiler et ah, 2001). 

Figure 11,4. ExPASy Proteomic tools. ExPASy server provides various tools for proteomic analysis which can be accessed from ExPASy Proteomic tools. These tools (locals or hyperlinks) include Protein identification and characterization, Translation from DNA sequences to protein sequences. Similarity searches, Pattern and profile searches, Post-translational modification prediction, Primary structure analysis, Secondary structure prediction, Tertiary structure inference, Transmembrane region detection, and Sequence alignment.

Knowledge of protein primary sequence, quantities, posttranslational modifications (PTMs), structures, protein-protein (P-P) interactions, cellular spatial relationships, and functions are seven important attributes (see Table 4.2) needed for comprehensive protein expression analysis. It is this multifold and complex nature of protein attributes that has spawned the development of so different many proteomic technologies. Some of these challenges in proteomic analysis include defining the identities and quantities of an entire proteome in a particular spatial location (i.e., serum, liver mitochondria, brain), the existence of multiple protein forms and complexes, the evolving structural and functional annotations of the human and rodent... 

The study of protein structure, function, quantity, and interactions during maturation and progression of disease is referred to as proteomics. Analytical approaches that use a combination of two-dimensional (2-D) gel electrophoresis for protein separation and MS analysis for protein identification followed by database searches is a widely practiced proteomics strategy.The tryptic peptides extracted from gels are analyzed by MALDI-TOF MS and microcolunm or capillary LC tandem mass spectrometry (MS/MS) techniques. Typically, the MALDI-TOF MS techniques are used to quickly identify peptide fragments and confirm the presence of known proteins. Nano-scale capillary LC/MS/MS techniques (using 50-100 pm diameter columns, operating at flow rates of 20-500 nL/min) are... 

Both these methods are time consuming and have their own technical difficulties. In addition, the structure of each protein must be solved individually through a laborious process. Thus, these are not high-throughput methods suitable for proteomic analysis. The computational approaches are more suitable in proteomics for determining the 3D protein structures from their primary structure, i.e., their amino acid sequence. 

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