Protein purification, basic steps

The best purification procedure is one that yields a maximum amount ol the purified protein in a minimum amount of time. The following discussion outlines the basic steps that must be considered in order to develop a protein purification scheme (Table E4.1). 

The thorough study of many biological systems has involved purification of one or more of the system components. Unfortunately, there are a large number of misconceptions about the manner in which to develop a successful purification scheme. One systematic way to develop such a scheme is discussed here using proteins as a model, but the same considerations, perhaps in different terms, would apply to any biologically significant molecule. The five basic steps of purification are as follows ... 

The field of proteome analysis is challenging because of the complexity of any given proteome, the broad dynamic range of protein abundance, the existence of mnltiple protein forms dne to PTMs, proteolysis and splice variants, and the dif-flcnlty of quantification of absolute protein content. Some of these challenges can be met by the use of complementary approaches to proteomics. In general, four basic steps are involved in proteome analysis (1) sample preparation, (2) separation and purification, (3) mass spectrometry analysis, and (4) database search. 

Although a few general guidelines exist, the design of such mentioned extraction schemes is still a trial-and-error proposition. Consequently, more basic information on the nature of the lipid -protein - surfactant interactions is still required. It should also be noted that in most instances, the micellar "extraction" step is merely the prelude to further fractionation (usually by electrophoretic, column or hydrophobic chromatographic techniques) and purification of the desired biological components (402-404). 

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