Expression and purification of recombinant proteins in E. coli

To ease the subsequent purification, the protein may be produced as a fusion protein, by inserting the open reading frame in frame with an affinity tag (see LaVallie and McCoy13 for a review). Among the most commonly used affinity tags are the following the Schistosoma japonicum glutathione S-transferase (GST) protein 

When producing proteins aimed for protein footprinting experiments (see Section 4.5) the purity, rather than the quantity of the expressed protein, is critical. By positioning the affinity-tag and the phosphorylation site at opposite ends of the introduced cDNA, only full-length protein becomes radiolabelled after purification14 (see Table 4.4 in Chapter 4). 

Proteins expressed in bacteria lack post-translational modifications such as phosphorylations and glycosylations. If these are crucial for function, expression in eukaryotic cells is a better choice. Procedures for expression of recombinant proteins in baculovirus or vaccinia virus can be found elsewhere.1516 

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