The application of DOSY NMR and molecular dynamics simulations to explore the mechanism(s) of micelle binding of antimicrobial peptides containing unnatural amino acids

Abstract

Anionic and zwitterionic micelles are often used as simple models for the lipids found in bacterial and mammalian cell membranes to investigate antimicrobial peptide-lipid interactions. In our laboratory we have employed a variety of 1D, 2D, and diffusion ordered (DOSY) NMR experiments to investigate the interactions of antimicrobial peptides containing unnatural amino acids with SDS and DPC micelles. Complete assignment of the proton spectra of these peptides is prohibited by the incorporation of a high percentage of unnatural amino acids which don't contain amide protons into the backbone. However preliminary assignment of the TOCSY spectra of compound 23 in the presence of both micelles indicated multiple conformers are present as a result of binding to these micelles. Chemical Shift Indexing agreed with previously collected CD spectra that indicated on binding to SDS micelles compound 23 adopts a mixture of α-helical structures and on binding to DPC micelles this peptide adopts a mixture of helical and β-turn/sheet like structures. DOSY NMR experiments also indicated that the total positive charge and the relative placement of that charge at the N-terminus or C-terminus are important in determining the mole fraction of the peptide that will bind to the different micelles. DOSY and (1) H-NMR experiments indicated that the length of Spacer #1 plays a major role in defining the binding conformation of these analogs with SDS micelles. Results obtained from molecular simulations studies of the binding of compounds 23 and 36 with SDS micelles were consistent with the observed NMR results.

Design and structural characterisation of monomeric water-soluble α-helix and β-hairpin peptides: State-of-the-art

“…In any case, spectral invariability upon dilution always discards the presence of non-specific aggregation. The monomeric or multimeric state of peptides can be confirmed by analytical ultracentrifugation [9][10][11], and by NMR diffusion ordered spectroscopy (DOSY) [12][13][14][15].…”

Effect of Liposome Surface Charge and Peptide Side Chain Charge Density on Antimicrobial Peptide-Membrane Binding as Determined by Circular Dichroism

“…Since the binding of a peptide to a SUVs effectively changes the molecular size of the peptide, binding interactions can be monitored by observing changes in the peptide's diffusion coefficient [40][41][42]55,[58][59][60][61][62][63][64][65]. When interpreting diffusion data it must be remembered that the binding of a peptide to the surface of a SUVs is a steady state equilibrium process between the SUV-bound peptide and the monomeric species in solution and the observed diffusion coefficient is the weight average of the two species [42,50,[55][56][57]59,60,65,66]. This equilibrium process is fast on the NMR time scale [59,60].…”

“…The common assumption is that the diffusion coefficient of a molecule is directly related to its' molecular weight. The diffusion coefficient is related to the hydrodynamic volume of the molecule not its molecular weight [66,67]. The interpretation of changes in the diffusion coefficient of a molecule must take into consideration changes in molecular volume that result from major conformational changes (changes from a globular to helical structure for example) in the molecule.…”

NMR diffusion: an update

“…Continuing with some further examples, DOSY NMR experiments have been employed to investigate the interactions of antimicrobial peptides containing synthetic amino acids with micelles (Clark et al, 2013), and have helped in the development of biomimetic dye polymers for photonics, based on Zn chlorin and poly(4-vinylpyridine), in that diffusionordered NMR studies have revealed that a coordinative interaction exists between the Zn chlorin and the pyridine polymer (Pale et al, 2013).…”