Advanced Protein Sequencing Techniques

In parallel with these advances in mRNA detection and assessment, significant advances in the detection, characterization, and quantitation of proteins have been made. This is not only a result of the rapidly expanding database of protein sequences but also of the extraordinary advances in sensitivity of mass spectrometry techniques. It is proposed, with some real sense of expected success, that protein characterization at the whole genome level will be forthcoming (Domon and Aebersold, 2006). 

Technological advances often lead theory and, for some time, studies of DNA and genomes have reigned as evidenced by Nobel Prizes and millions of dollars in ventnre capital. In more recent years, interest in proteins has grown as researchers realized that DNA and RNA are only one half of life s equation. This renewed interest in proteins has taken a new name, structural genomics, and is driven by new techniqnes in protein chemistry and major advances with older techniques. Extensive genetic and biochemical efforts in the last decades have led to a detailed description of many of the molecular pathways that constitute normal and aberrant cellnlar fnnctions. The recent completion of the DNA sequences of a number of genomes will lead to an unprecedented description of cellular processes at the molecular level. 

It is assumed that the limited proteolysis phenomenon derives from the fact that a specific polypeptide chain segment of the compact, folded protein substrate is exposed and flexible so that it can fit the active site of the appropriate peptidase for an efficient and selective limited hydrolysis. There is no doubt that enhanced chain flexibility or segment mobility is the key feature of the site of peptide bond hydrolysis demonstrated by a clear-cut correlation between sites of proteolytic attack and sites of enhanced chain flexibility. The present availability of automatic, efficient and sensitive techniques of protein sequencing and, particularly, the recent dramatic advances of mass spectrometry 361 in the analysis of peptides and proteins, allows a more systematic use of the limited proteolysis approach as a simple first step in the elucidation of structure-dynamics-function relationships for novel proteins which are only available in minute amounts. 

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