Expression of recombinant staphylokinase, a fibrin-specific plasminogen activator of bacterial origin, in potato (Solanum tuberosum L.) plants

Abstract

One of the most dynamically developing sectors of green biotechnology is molecular farming using transgenic plants as natural bioreactors for the large scale production of recombinant proteins with biopharmaceutical and therapeutic values. Such properties are characteristic of certain proteins of bacterial origin, including staphylokinase. For many years, work has been carried out on the use of this protein in thrombolytic therapy. In this study, transgenic Solanum tuberosum plants expressing a CaMV::sak-mgpf-gusA gene fusion, were obtained. AGL1 A. tumefaciens strain was used in the process of transformation. The presence of the staphylokinase gene was confirmed by PCR in 22.5% of the investigated plants. The expression of the fusion transgene was detected using the β-glucuronidase activity assay in 32 putative transgenic plants. Furthermore, on the basis of the GUS histochemical reaction, the transgene expression pattern had a strong, constitutive character in seven of the transformants. The polyacrylamide gel electrophoresis of a protein extract from the SAK/PCR-positive plants, revealed the presence of a119 kDa protein that corresponds to that of the fusion protein SAK-mGFP-GUSA. Western blot analysis, using an antibody against staphylokinase, showed the presence of the staphylokinase domain in the 119 kDa protein in six analyzed transformants. However, the enzymatic test revealed amidolytic activity characteristic of staphylokinase in the protein extract of only one plant. This is the first report on a Solanum tuberosum plant producing a recombinant staphylokinase protein, a plasminogen activator of bacterial origin.

The pharmaceutics from the foreign empire: the molecular pharming of the prokaryotic staphylokinase in Arabidopsis thaliana plants

“…So far, our attempts to produce SAK in plant tissues with the use of the CaMV35S promoter had limited success. Thus, the described research project makes a continuation of the studies over the recombinant staphylokinase production in plants conducted in our Department (Gerszberg et al 2012 ). However, in the light of our past results, we decided to change the expression cassette components.…”

Tomato tolerance to abiotic stress: a review of most often engineered target sequences

“…Over the past two decades, numerous techniques were used to introduce foreign genes into both mono-and dicotyledonous plants, such as rice, potato, soya bean, tomato or common bean (Sahoo et al 2011;Gerszberg et al 2012;Hnatuszko-Konka et al 2014). The first protocol for the genetic transformation of Solanum lycopersicum was reported in the 1980s and since then a significant progress in this field has been made (McCormick et al 1986).…”

Compendium on Food Crop Plants as a Platform for Pharmaceutical Protein Production

“…At this stage of cassette design, a decision on the selection and monitoring of transformant must be made as well. Therefore, selection or reporter genes (e.g., gus , gfp , kan ) should be present within the expression construct [ 34 , 35 , 36 ].…”