By O. Zerbe
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Additional info for BioNMR in Drug Research
5 Alternatives to E. coli Expression Systems Here, we describe the various alternatives to the use of E. coli expression hosts for heterologous gene expression. Advantages and disadvantages for the different expression systems are discussed, and practical aspects of expression technologies are also described. The feasibility of isotope labeling of recombinantly expressed proteins and their potential use for NMR is also discussed, since the costs and quantities of recombinant proteins produced depend on the system being used.
More dramatically, expression studies of fusion constructs between the Bop gene and mammalian GPCRs (G protein-coupled receptors, human muscarinic M1 receptor, platelet-activating receptor and angiotensin-1 receptor) failed to detect fusion protein expression detected by Western blotting . In this case, coding region swaps between Bop and GPCRs improved RNA yields and resulted in detectable levels of Ste2 receptor. These results suggest that H. salinarum can be considered as a potentially interesting alternative.
5 A), analogous to RNA splicing [57, 58]. This unique enzymatic process has been used in various biochemical and biotechnological applications such as protein purification, protein ligation, backbone cyclization and C-terminal modifications. In particular, protein ligation using inteins has opened up a new way for the production of segmentally labeled proteins, thereby reducing the complexity of NMR spectra. For larger proteins or multimeric proteins, it may be necessary to combine selective labeling and segmental isotope-labeling approaches.
BioNMR in Drug Research by O. Zerbe