Журнал аналитических и биоаналитических методов

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Targeted Protein Delivery Using Modified Anthrax Toxin Protective Antigen

Dr. Verdurmen Richa

Background: It would be highly desired to develop effective strategies for trans locating tailored binding proteins to the cytosol since they might target flat and hydrophobic protein-protein interfaces, which would expand the amount of the druggable proteome. Two levels of specificity would be gained, one for the cell type and the other for the cytosolic target, if this could be done in a way that was dependent on a cell surface receptor. Such mechanisms have naturally developed in bacterial protein poisons. Protective antigen (PA), a translocation unit that forms pores, and a distinct protein payload make up anthrax toxin. By combining a chosen ankyrin repeat protein with PA, it is possible to engineer PA to abolish binding to its own receptor and instead bind to a desired receptor, leading to uptake in different cell types.

Results: The amount of redirected PA that can be administered and subsequently the amount of delivered payload are both constrained by prepore-to-pore conversion of redirected PA that already takes place at the cell surface. We proposed that the deficiency of a stabilising interaction with the wild-type PA receptor is the cause. Now, PA has been redesigned to include the CMG2 anthrax receptor binding domain followed by a DARPin that binds to the desired receptor. This construct may be supplied at considerably higher concentrations without causing toxicity, undergoes prepore-to-pore conversion only in late endosomes, and is thus stable. As a result, it delivers substantially larger payload levels to the cytosol.

Conclusion: We think that this reengineered system represents a significant advancement in the quest for effective protein delivery to the cytosol in specific cells.