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Maximized Autotransporter-Mediated Expression (MATE) for Surface Display and
Secretion of Recombinant Proteins in Escherichia coli   

Shanna Sichwart1, Iasson E.P. Tozakidis1,2, Mark Teese1,3 and Joachim Jose1,2

1Institute of Pharmaceutical and Medicinal Chemistry, PharmaCampus, Westphalian Wilhelms-University,
 Corrensstraße 48, DE-48149 Münster, Germany
2The NRW Graduate School of Chemistry, Wilhelm-Klemm-Straße 10, DE-48149 Münster, Germany
3Present address: Technical University Munich, Weihenstephaner Berg 3, DE-85354 Freising, Germany




Article history:
Received July 8, 2014
Accepted March 27, 2015



Key words
surface display, secretion, autotransporter, mCherry, OmpT, EhaA, MATE




Summary
A new optimized system for the surface display and secretion of recombinant proteins is described, termed MATE (maximized autotransporter-mediated expression). It is basedon an artificial gene consisting of the coding region for the signal peptide of CtxB, a multiple cloning site for passenger gene insertion, flanked by coding sequences for linear epitopes for monoclonal antibodies and OmpT, and factor Xa protease cleavage sites followed by a codon-optimized DNA sequence of the linker and the β-barrel of the type V autotransporter EhaA from Escherichia coli under control of an IPTG-inducible T5 promoter. The MATE system enabled the continuous secretion of recombinant passenger mCherry via OmpT-mediated cleavage, using native OmpT protease activity in E. coli when grown at 37 °C. It is the first example to show that native OmpT activity is sufficient to facilitate the secretion of a correctly folded target protein in preparative amounts obtaining 240 μg of purified mCherry from 800 mL of crude culture supernatant. Because the release of mCherry was achieved by a simple transfer of the encoding plasmid from an OmpT-negative to an OmpT-positive strain, it bears the option to use surface display for screening purposes and secretion for production of the selected variant. A single plasmid could therefore be used for continuous secretion in OmpT-positive strains or surface display in OmpT-negative strains. In conclusion, the MATE system appears to be a versatile tool for the surface display and for the secretion of target proteins in E. coli.







*Corresponding author:   email3  joachim.jose@uni-muenster.de                                    
                                       tel3  +49 (0)251 8332 200
                                       fax2  +49 (0)251 8332 211  

 


 


 





 

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