Specific uptake and interactions of peptide nucleic acid derivatives with biomimetic membranes

Abstract

There is a growing interest in understanding the uptake mechanism of metal-containing peptide nucleic acid (PNA) bioconjugates into living cells. In this study, quartz crystal microbalance with dissipation monitoring (QCM-D) has been used to explore the membrane specific uptake and interactions of PNA/peptide/Ru(II) conjugates. For all lipid compositions, the unmodified PNA oligomer and its Ru(II) conjugate were found to traverse freely across the membrane in a trans-membrane manner, causing no significant changes in the membrane structure. The nuclear localised signal peptide (NLS) conjugated sequences showed membrane specific activities. In model mammalian and bacterial-mimetic membranes, rapid trans-membrane insertion was observed followed by a concentration dependent disruption and irreversible structural changes to the membrane system. The variations in the magnitude of the structural changes and in their tendency to facilitate disruption are ascribed to hydrophobicity, the cationic charge introduced on modification of the original PNA backbone as well as the physical state of the model membrane used. There is a growing interest in understanding the uptake mechanism of metal-containing peptide nucleic acid (PNA) bioconjugates into living cells. In this study, Quartz Crystal Microbalance with Dissipation monitoring (QCM-D) has been used to explore the membrane specific uptake and interactions of PNA/peptide/Ru(II) conjugates. For all lipid compositions, the unmodified PNA oligomer and its Ru(II) conjugate were found to traverse freely across the membrane in a trans-membrane manner, causing no significant changes in 10 membrane structure. The Nuclear Localised Signal Peptide (NLS) conjugated sequences showed membrane specific activities. In model mammalian and bacterial-mimetic membranes, rapid trans-membrane insertion was observed followed by a concentration dependent disruption and irreversible structural changes to the membrane system. The variations in the magnitude of the structural changes and in their tendency to facilitate disruption are ascribed to hydrophobicity, the cationic charge introduced on modification of the original PNA 15 backbone as well as the physical state of the model membrane used.

Real-time examination of aminoglycoside activity towards bacterial mimetic membranes using Quartz Crystal Microbalance with Dissipation monitoring (QCM-D)

“…We have previously shown that these traces provide a unique "fingerprint" characteristic of the mode of interaction of a small molecule or biomolecule with the supported phospholipid bilayer-modified sensor [51,53,54,65]. The Δf-ΔD fingerprints for both kanamycin A and neomycin B are shown in Fig.…”

“…QCM-D experiments are also able to provide depth profiling via analysis of frequency changes at different harmonics of the QCM-D sensor [51,[53][54][55]. Each harmonic probes a defined distance away from the surface of the sensor.…”

“…The sensors were treated with mercaptopropionic acid to create a selfassembled carboxylate monolayer prior to the introduction of the liposomes as described in the Supporting Information. All QCM-D experiments were conducted at 19.10 ± 0.05°C [52], in triplicate, using established experimental protocols also used for peptidesmembrane interactions (see Supporting Information for details) [51,[53][54][55]. Concentration studies were carried out using 1, 10 and 15 μM solutions of kanamycin A and neomycin B in high-salt PBS.…”

“…A lipid composition of DMPC/DMPG (4:1 v/v) was used for liposome preparation (see Supporting Information for details), following our previously published procedure [50,51]. The Quartz Crystal Microbalance with Dissipation Monitoring (QCM-D) measurements were performed using E4 system with flow cells (Q-Sense, Västra Frölunda, Sweden).…”

Di-heterometalation of thiol-functionalized peptide nucleic acids

“…45 Over the past few years, research in our groups has focused on the design and evaluation of metal-PNA bioconjugates for multimodal biosensing, nuclear medicine, molecular and cell biology applications. [46][47][48][49][50][51] Toward this end, targeting the development of robust direct (solid-phase) and indirect (solution phase) ways for the preparation of multi-metallic PNA conjugates, we recently reported the first di-hetero-organometallic-containing PNA the cyclopentadienyl moiety at ca. 4.64 ppm.…”