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Membrane Crossing and Membranotropic Activity of Cell- Penetrating Peptides: Dangerous Liaisons?

Acc. Chem. Research2017


In this Account, we focus on cationic cell-penetrating peptides (CPPs) and the way they cross cell membranes. We summarize the history of this fi eld that emerged around 20 years ago. CPPs were indeed fi rst identifi ed as protein-transduction domains from the human immunodefi ciency virus (HIV) TAT protein and the Antennapedia homeoprotein, a transcription factor from Drosophila. We highlight our contribution to the elucidation of CPP internalization pathways, in particular translocation, which implies perturbation and reorganization of the lipid bilayer, and endocytosis depending on sulfated glycosaminoglycans. We show a particular role of Trp (indole side chain) and Arg (guanidinium side chain), which are essential amino acids for CPP internalization. Interactions with the cell-surface are not only Coulombic; H-bonds and hydrophobic interactions contribute also significantly to CPP entry. The capacity of CPPs to cross cell membrane is not related to their strength of membrane binding. Finally, we present optimized methods based on mass spectrometry and fluorescence spectroscopy that allow unequivocal quantification of CPPs inside cells or bound to the outer leafl et of the membrane, and discuss some limitations of the technique of flow cytometry that we have recently highlighted.

Structure and dynamics of an intrinsically disordered protein region that partially folds upon binding by chemical-exchange NMR

Journal of the American Chemical Society, Volume 139, p. 12219 - 12227, 2017


Many intrinsically disordered proteins (IDPs) and protein regions (IDRs) engage in transient, yet specific, interactions with a variety of protein partners. Often, if not always, interactions with a protein partner lead to partial folding of the IDR. Characterizing the conformational space of such complexes is challenging: in solution-state NMR, signals of the IDR in the interacting region become broad, weak and often invisible; while X-ray crystallography only provides information on fully ordered regions. There is thus a need for a simple method to characterize both fully and partially ordered regions in the bound state of IDPs. Here, we introduce an approach based on monitoring chemical exchange by NMR to investigate the state of an IDR that folds upon binding through the observation of the free state of the protein. Structural constraints for the bound state are obtained from chemical shifts and site-specific dynamics of the bound state are characterized by relaxation rates. The conformation of the interacting part of the IDR was determined and subsequently docked onto the structure of the folded partner. We apply the method to investigate the interaction between the disordered C-terminal region of Artemis and the DNA binding domain of Ligase IV. We show that we can accurately reproduce the structure of the core of the complex determined by X-ray crystallography and identify a broader interface. The method is widely applicable to the biophysical investigation of complexes of disordered proteins and folded proteins.

Re(I) carbonyl complexes: Multimodal platforms for inorganic chemical biology

Coordination Chemistry Reviews 351 (2017) 172–188


Bio-imaging, by enabling the visualization of biomolecules of interest, has proved to be highly informative in the study of biological processes. Although fluorescence microscopy is probably one of the most used techniques, alternative methods of imaging, providing complementary information, are emerging. In this context, metal complexes represent valuable platforms for multimodal imaging, since they may combine interesting spectroscopic features and biologically relevant functionalization on a single molecular core. In particular, d6 low-spin rhenium tri-carbonyl complexes display unique luminescence and vibrational properties, and can be readily functionalized. Here we review their applications and potential as probes or drugs relying on their photophysical properties, before focusing on their use as multimodal probes for the labelling and imaging of peptides and proteins.



Exploiting Benzophenone Photoreactivity to Probe the Phospholipid Environment and Insertion Depth of the Cell-Penetrating Peptide Penetratin in Model Membranes

Angew Chem Int Ed. 2017 May 9


Penetratin (RQIKIWFQNRRMKWKK) enters cells by different mechanisms, including membrane translocation, implying that the peptide interacts with the lipid bilayer. Penetratin also crosses the membrane of artificial vesicles depending on their phospholipid content. To evaluate the phospholipid preference of Penetratin, as the first step of translocation, we have exploited the benzophenone triplet kinetics of hydrogen abstraction, slower for secondary than for allylic hydrogens. Using multilamellar vesicles (MLVs) of various phospholipid content, we have identified and characterized the crosslinked products by MALDI-TOF mass spectrometry. Penetratin shows a preference for negatively charged (vs zwitterionic) polar heads, for unsaturated (vs saturated) and short (vs long) saturated phospholipids. Our study highlights the potential of using benzophenone to probe the environment and insertion depth of membranotropic peptides in membranes.

Interview Geoffrey BODENHAUSEN - Comprendre la Résonance magnétique nucléaire

La résonance magnétique nucléaire (RMN) est un outil analytique universel de détermination structurale et dynamique de la matière au sens large : produits naturels, substances synthétiques, macromolécules biologiques… jusqu’au corps humain tout entier grâce à l’imagerie par résonance magnétique (IRM).