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Fluorescence Study of the Interaction between the TAT‐PTD Peptide and the Lipid Bilayer
Author(s) -
Tiriveedhi Venkataswarup,
Butko Peter
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.6.a980-c
Subject(s) - chemistry , fluorescence anisotropy , lipid bilayer , diphenylhexatriene , membrane fluidity , membrane , fluorescence , bilayer , biophysics , tryptophan , pyrene , biochemistry , organic chemistry , physics , biology , amino acid , quantum mechanics
The mechanism of internalization of cationic peptides into the cells is unknown. We used fluorescence spectroscopy to study the interaction between Protein Transduction Domain of the HIV‐1 TAT protein (TAT‐PTD; residues 47–60 of TAT, fluorescently labeled with tryptophan) and the lipid bilayer in the form of small unilamellar lipid vesicles (SUV). The TAT‐PTD tryptophan exhibited a decrease in fluorescence intensity and increase in anisotropy upon the interaction with SUV, which was proportional with the negative charge density in the membrane. Kinetic analysis of the interaction showed two apparent dissociation constants. Kd1 was independent of the negative charge density and accounted for 24% of the interaction, whereas Kd2, contributing 76% to the interaction, decreased linearly with the density of negative charge in the membrane, suggesting an electrostatic nature of the latter interaction. The former could not be inhibited by high salt, suggesting its van der Waals or hydrophobic nature. TAT‐PTD did not dissipate membrane potential (165 mV, negative inside), nor did it affect fluorescence anisotropy of the membrane fluorescence probes TMA‐DPH or DPH, indicating preserved membrane integrity upon TAT‐PTD binding. When the pyrene‐labeled phosphatidyl glycerol was included in the membrane, TAT‐PTD induced pyrene excimer formation, but only at the temperature above the melting point of the lipid. Thus, the two prerequisites for a strong binding of TAT‐PTD to the lipid membrane are the presence of the negative charge and the liquid crystalline phase of the lipid bilayer.