EDDIE fusion proteins: Triggering autoproteolytic cleavage

Efficient biosynthesis of a Cecropin A-melittin mutant in Bacillus subtilis WB700

“…This function allows it to release its fusion partner from the C-terminal end of the autoprotease by self-cleavage, resulting in a target protein with an authentic N-terminus21. Furthermore, its tailor-made mutant (EDDIE) shows improved solubility and faster cleavage than its parent and has already been applied to improve heterologous expression by overcoming challenges such as inefficient cleavage and proteolytic degradation22. EDDIE has been applied to produce CM4 in host E. coli strains and to protect host cells from CM4-induced damage; however, the fusion protein was expressed in inclusion bodies, and a complicated procedure was required to obtain the active product23.…”

High-level soluble expression of hIGF-1 fusion protein in recombinant Escherichia coli

“…Therefore, several fusion tag strategies have been applied to enhance expression [10]. In our previous work, several soluble human defensins have been overexpressed in E. coli as TrxAfusion protein, and the bioactive defensins could be achieved after the enterokinase digestion of fusion protein [11].…”

Self-cleaving fusion tags for recombinant protein production

“…The use of this SpeI site thus allows insertion of the target gene without introduction of any non-native residues (Achmüller et al 2007;Cheng et al 2010). Although the N pro fusion system allows protein with an authentic N-terminus to be produced, the first amino acid of the target protein has a major impact on the cleavage rate and an N-terminal proline fully blocks the cleavage (Achmüller et al 2007;Ueberbacher et al 2009). Furthermore, the overall reaction can be predominantly influenced by the target protein.…”

“…Due to the above reasons, the N pro fusion system is particular appropriate for the production of small peptides in which case the property of the fusion protein is mainly determined by N pro . For N pro fusions containing small target peptides, refolding and cleavage yield is independent of initial protein concentration (Ueberbacher et al 2009). Schmoeger et al (2010) have recently demonstrated that matrixassisted refolding is superior to conventional dilution refolding regarding cleavage rate and yield.…”