Abstract
Since the introduction of recombinant protein expression in the second half of the 1970s, the growth of the biopharmaceutical field has been rapid and protein therapeutics has come to the foreground. Biophysical and structural characterisation of recombinant proteins is the essential prerequisite for their successful development and commercialisation as therapeutics. Despite the challenges, including low protein solubility and inclusion body formation, prokaryotic host systems and particularly Escherichia coli, remain the system of choice for the initial attempt of production of previously unexpressed proteins. Several different approaches have been adopted, including optimisation of growth conditions, expression in the periplasmic space of the bacterial host or co-expression of molecular chaperones, to assist correct protein folding. A very commonly employed approach is also the use of protein fusion tags that enhance protein solubility. Here, a range of experimentally tested peptide tags, which present specific advantages compared to protein fusion tags and the concluding remarks of these experiments are reviewed. Finally, a concept to design solubility-enhancing peptide tags based on a protein’s pI is suggested.
Highlights
Since the first successful attempt at recombinant production of the human peptide hormone Somatostatin in Escherichia coli in 1976 [1], protein therapeutics have come a long way
The average charge of a protein at a certain pH value depends on the pI, which can be calculated from the pKa values of the side chains of the residues that are ionised
It becomes obvious that the choice of the most appropriate solubility-enhancing tag depends on the individual protein and requires careful design; generalisation should be avoided [64]
Summary
Since the first successful attempt at recombinant production of the human peptide hormone Somatostatin in Escherichia coli in 1976 [1], protein therapeutics have come a long way. The biochemical characterisation of proteins is of utmost significance and a prerequisite prior to their commercialisation This process requires sufficient amounts of protein, which can be generated with the use of recombinant technology. E. coli is characterised by rapid growth at a low cost, with a cell doubling time of approximately 20 min [6]. This fast growth rate, in combination with the range of plasmids and safe, compatible strains that can be exploited, which can be flexibly tailored to the individual needs of recombinant production, makes this organism the ideal host [7]
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