Comparison of microbial and transient expression (tobacco plants and plant‐cell packs) for the production and purification of the anticancer mistletoe lectin viscumin

<jats:title>Abstract</jats:title><jats:p>Cancer is the leading cause of death in industrialized countries. Cancer therapy often involves monoclonal antibodies or small‐molecule drugs, but carbohydrate‐binding lectins such as mistletoe (<jats:italic>Viscum album</jats:italic>) viscumin offer a potential alternative treatment strategy. Viscumin is toxic in mammalian cells, ruling them out as an efficient production system, and it forms inclusion bodies in <jats:italic>Escherichia coli</jats:italic> such that purification requires complex and lengthy refolding steps. We therefore investigated the transient expression of viscumin in intact <jats:italic>Nicotiana benthamiana</jats:italic> plants and <jats:italic>Nicotiana tabacum</jats:italic> Bright Yellow 2 plant‐cell packs (PCPs), comparing a full‐length viscumin gene construct to separate constructs for the A and B chains. As determined by capillary electrophoresis the maximum yield of purified heterodimeric viscumin in <jats:italic>N. benthamiana</jats:italic> was ~7 mg/kg fresh biomass with the full‐length construct. The yield was about 50% higher in PCPs but reduced 10‐fold when coexpressing A and B chains as individual polypeptides. Using a single‐step lactosyl‐Sepharose affinity resin, we purified viscumin to ~54%. The absence of refolding steps resulted in estimated cost savings of more than 80% when transient expression in tobacco was compared with <jats:italic>E. coli</jats:italic>. Furthermore, the plant‐derived product was ~3‐fold more toxic than the bacterially produced counterpart. We conclude that plants offer a suitable alternative for the production of complex biopharmaceutical proteins that are toxic to mammalian cells and that form inclusion bodies in bacteria.</jats:p>