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Label-free probe of HIV-1 TAT peptide binding to mimetic membranes

Author:
Rao, Yi, Kwok, Sheldon J. J., Lombardi, Julien, Turro, Nicholas J., Eisenthal, Kenneth B.
Source:
Proceedings of the National Academy of Sciences of the United States of America 2014 v.111 no.35 pp. 12684-12688
ISSN:
0027-8424
Subject:
Human immunodeficiency virus 1, amino acid sequences, cell membranes, dissociation, equations, phospholipids
Abstract:
The transacting activator of transduction (TAT) protein plays a key role in the progression of AIDS. Studies have shown that a +8 charged sequence of amino acids in the protein, called the TAT peptide, enables the TAT protein to penetrate cell membranes. To probe mechanisms of binding and translocation of the TAT peptide into the cell, investigators have used phospholipid liposomes as cell membrane mimics. We have used the method of surface potential sensitive second harmonic generation (SHG), which is a label-free and interface-selective method, to study the binding of TAT to anionic 1-palmitoyl-2-oleoyl- sn -glycero-3-phospho-1′- rac -glycerol (POPG) and neutral 1-palmitoyl-2-oleoyl- sn -glycero-3-phosphocholine (POPC) liposomes. It is the SHG sensitivity to the electrostatic field generated by a charged interface that enabled us to obtain the interfacial electrostatic potential. SHG together with the Poisson–Boltzmann equation yielded the dependence of the surface potential on the density of adsorbed TAT. We obtained the dissociation constants K d for TAT binding to POPC and POPG liposomes and the maximum number of TATs that can bind to a given liposome surface. For POPC K d was found to be 7.5 ± 2 μM, and for POPG K d was 29.0 ± 4.0 μM. As TAT was added to the liposome solution the POPC surface potential changed from 0 mV to +37 mV, and for POPG it changed from −57 mV to −37 mV. A numerical calculation of K d, which included all terms obtained from application of the Poisson–Boltzmann equation to the TAT liposome SHG data, was shown to be in good agreement with an approximated solution.
Agid:
1787096