Influence of protein transduction domains on target-specific chimeric proteins.
Direct targeting to the cytoplasm and nucleus using protein transduction domains (PTD) has been described to be efficient but non-cell-type-specific, and only has clinical relevance when the molecule is active exclusively in the diseased cell. The use of PTDs is an attractive mechanism to improve drug delivery. In this work, we designed recombinant proteins that contain epidermal growth factor as ligand to render uptake target cell-specific. We evaluated the potential of several PTDs to induce the cytosolic uptake of the catalytic domain of diphtheria toxin by measuring cytotoxicity. Although PTD-dependent membrane transfer is very low, the proteins exhibited concentration-dependent cytotoxic activity. Higher binding at 4 degrees C compared to 37 degrees C suggests that uptake by the PTDs MTS and TLM occurs via an endocytic pathway. Non-specific binding is predominantly a function of the PTD and greatly increases by substitution of a non-polar glycine with a negatively charged glutamate in the PTD HA2.
Bachran
Christopher
C
Institut für Klinische Chemie und Pathobiochemie, Charité-Universitätsmedizin Berlin, Campus Benjamin Franklin, Germany.
Heisler
Iring
I
Fuchs
Hendrik
H
Sutherland
Mark
M
D000595:Amino Acid Sequence; D002478:Cells, Cultured; D016503:Drug Delivery Systems; D018698:Glutamic Acid / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D011485:Protein Binding; D017434:Protein Structure, Tertiary / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013696:Temperature; D014161:Transduction, Genetic
D011993:Recombinant Fusion Proteins; D018698:Glutamic Acid