Mechanistic insight into how polycations disrupt and cross cell membranes is needed for understanding and controlling polycation-membrane interactions, yet such information is surprisingly difficult to obtain at the molecular level. We use second harmonic and vibrational sum frequency generation spectroscopies along with quartz crystal microbalance with dissipation monitoring and computer simulations to quantify the interaction of poly(allylamine) hydrochloride (PAH) and its monomeric precursor allylamine hydrochloride (AH) with lipid bilayers. We find PAH adsorption to be reversible and nondisruptive to the bilayer under the conditions of our experiments. With an observed free adsorption energy of -52.7 ± 0.6 kJ/mol, PAH adsorption was found to be surprisingly less favorable relative to AH (-14.6 ± 0.4 kJ/mol) when considering a simple additive model. By experimentally quantifying the number of adsorbates and the average amount of charge carried by each adsorbate, we find that the PAH is associated with only 70% of the positive charges it could hold while the AH remains mostly charged while attached to the membrane. Simulations indicate that PAH pulls in condensed counterions from solution to avoid charge-repulsion along its backbone and with other PAH molecules to attach to, and completely cover, the bilayer surface. In addition, computations indicate that the amine groups shift their pK values due to the confined environment upon adsorption to the surface. Our results provide experimental constraints for theoretical calculations, which yield atomistic views of the structures that are formed when polycations interact with lipid membranes that will be important for predicting polycation-membrane interactions.

We report vibrational sum frequency generation (SFG) spectra in which the C-H stretches of lipid alkyl tails in fully hydrogenated single-and dual-component supported lipid bilayers are detected along with the O-H stretching continuum above the bilayer. As the salt concentration is increased from ~10 µM to 0.1 M, the SFG intensities in the O-H stretching region decrease by a factor of 2, consistent with significant absorptive-dispersive mixing between χ (2) and χ (3) contributions to the SFG signal generation process from charged interfaces.A method for estimating the surface potential from the second-order spectral lineshapes (in the OH stretching region) is presented and discussed in the context of choosing truly zero-potential reference states. Aided by atomistic simulations, we find that the strength and orientation distribution of the hydrogen bonds over the purely zwitterionic bilayers are largely invariant between sub-micromolar and hundreds of millimolar concentrations. However, specific Dogangun et al.Page 2 interactions between water molecules and lipid headgroups are observed upon replacing phosphocholine (PC) lipids with negatively charged phosphoglycerol (PG) lipids, which coincides with SFG signal intensity reductions in the 3100 cm -1 to 3200 cm -1 frequency region.The atomistic simulations show that this outcome is consistent with a small, albeit statistically significant, decrease in the number of water molecules adjacent to both the lipid phosphate and choline moieties per unit area, supporting the SFG observations. Ultimately, the ability to probe hydrogen-bond networks over lipid bilayers holds the promise of opening paths for understanding, controlling, and predicting specific and non-specific interactions between membranes and ions, small molecules, peptides, polycations, proteins, and coated and uncoated nanomaterials.I. Introduction. The structure of water over lipid membranes is of interest for a variety of reasons that are rooted in fundamental scientific interest and connect all the way to biological function and technological applications. 1-6 Specific questions pertain to whether there exist populations of interfacial water molecules that can undergo hydrogen-bond (H-bond) interactions with certain membrane constituents that can be strengthened or weakened with variations in ionic strength, or, as indicated by molecular dynamics simulations, 2 whether some population of water molecules exists that may interact specifically with certain lipid headgroups over others.While interface-specific vibrational spectroscopic approaches, particularly those that are based on sum frequency generation (SFG), are in principle well suited for probing water near membranes, this method has been largely limited to probing lipid monolayers 1, 7-16 chemically asymmetric bilayers, [17][18][19] or the use of D 2 O as opposed to H 2 O. 20-21 Indeed, the use of SFG spectroscopy for probing fully hydrogenated lipid bilayers is now just emerging. Part of the Dogangun et al. Page 3 reason for this relatively new ...

Quantifying the number of charges on peptides bound to interfaces requires reliable estimates of (i) surface coverage and (ii) surface charge, both of which are notoriously difficult parameters to obtain, especially at solid/water interfaces. Here, we report the thermodynamics and electrostatics governing the interactions of l-lysine and l-arginine octamers (Lys8 and Arg8) with supported lipid bilayers prepared.