Towards the improvement of a rhizosecretion-based recombinant protein production system: Developing protease-depleted lines of Arabidopsis thaliana.

Besides traditional production systems, such as bacteria, yeasts and mammal cells, plants can now be used to produce eukaryotic recombinant proteins. Their advantages as hosts for protein production include correct post-translational modifications, low cost of maintenance and no risk of contamination by human pathogens. Targeting heterologous proteins to the extracellular space is required for the correct folding of complex proteins and makes harvesting and purification easier. However, the quantity and the quality of recombinant proteins have been proved to be reduced by the action of endogenous co-secreted proteases.
In this study, we aimed at identifying active root-secreted (rhizosecreted) proteases in the model plant Arabidopsis thaliana. Their activity was assayed by in vitro degradation of a target protein (Bovine Serum Albumine, BSA) in a range of pH. The protease classes involved in BSA degradation were evaluated by inhibitor-based assays that revealed serine proteases as the major class involved in this degradation in any tested conditions. As a first step towards identification, and subsequent silencing, of the most active members of this class, rhizosecreted proteases are being analyzed by the “Activity-Based Protein Profiling” approach.