Binding proteins internalized by PTD-fused ligands allow the intracellular sequestration of selected targets by ligand exchange.
The targeted inactivation of intracellular molecules has important therapeutic potential. For this purpose, it could be envisioned to introduce specifically designed binding proteins into cells by covalent linkage to protein transduction domains (PTDs). However, stable linkage of a PTD to a cargo may affect its conformation and, hence, its binding property inside the cell. Here, we analyzed the ability of non-covalently linked PTDs to internalize the model binding proteins streptavidin (SA) and Strep-Tactin (ST). Notably, inside the cell, the PTD-Strep-tag II ligand used for internalization of SA was displaced by the model target biotin which exhibits a higher binding affinity for the same binding pocket. Thus, specifically designed binding proteins can be internalized into cells by non-covalent binding to a PTD and subsequently be used for capturing given intracellular target molecules by ligand exchange. Under therapeutic aspects, it could be envisioned to further develop such systems for the intracellular sequestration, and consequently, functional inactivation of pathologically relevant factors.
Moosmeier
Markus A
MA
Molecular Therapy of Virus-Associated Cancers (F065), German Cancer Research Center, D-69120 Heidelberg, Germany. markus.moosmeier@morphosys.com
Bulkescher
Julia
J
Hoppe-Seyler
Karin
K
Hoppe-Seyler
Felix
F
D001710:Biotin / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism*; D004705:Endocytosis; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006801:Humans; D042541:Intracellular Space / Q000378:metabolism*; D008024:Ligands; D008954:Models, Biological; D011485:Protein Binding; D055503:Protein Multimerization; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D019809:Streptavidin / Q000378:metabolism; D013696:Temperature; D054322:tat Gene Products, Human Immunodeficiency Virus / Q000378:metabolism
D002352:Carrier Proteins; D008024:Ligands; D011993:Recombinant Fusion Proteins; D054322:tat Gene Products, Human Immunodeficiency Virus; D001710:Biotin; D019809:Streptavidin