Direct measurement of membrane dipole field in complex model membranes via vibrational stark effect spectroscopy coupled with molecular dynamics simulations

Abstract

The heterogeneous composition of a biological membrane creates a complex electrostatic environment that regulates membrane structure and function. In this work, we investigated the magnitude of the membrane dipole field, Fd, located entirely within the low dielectric membrane interior as a function of membrane composition complexity. We directly measured Fd in vesicle model membrane composed of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) using vibrational Stark effect (VSE) shifts of nitrile oscillators systematically placed along the membrane interior coupled with extensive Molecular Dynamics (MD) simulations. We calculated the absolute magnitude of Fd in DMPC vesicles to be 8-11 MV/cm, at the high end of the range provided in literature. We increased the complexity of the membrane composition by intercalating cholesterol molecule at a wide range of concentration (0- 40 mol%) and found that cholesterol increased Fd at low concentration (~10 mol%), and decreased Fd at higher concentration (>10 mol%). This result, when compared to lipid bilayers containing a cholesterol derivative, 6-ketocholestanol (6-kc) that differs from cholesterol by only a ketone functional group, was strikingly different. Using the spectral line widths obtained from Fourier-transform infrared spectroscopy experiments and molecular dynamic simulations on model lipid-sterol bilayers, we propose that the membrane dipole field is greatly correlated to the local membrane structure and organization regulated by the sterols in the bilayer. We propose that at low concentrations, cholesterol increases dipole field by increasing packing density of disordered lipids, cholesterol and their associated hydrogen bonded water dipoles whereas at high concentrations, the sterol decreases the field by forming liquid ordered state enriched in cholesterol, thus spacing out phospholipids along with water dipoles. 6-kc, on the other hand, is homogeneously distributed and increases hydrogen bonding with water dipoles via two polar groups on its sterol ring, thus never promoting ordered domain and increasing the dipole field monotonously. We also investigated the translocation mechanism of positively, negatively and zwitterionic charged tryptophan molecules through a phospholipid bilayer using time dependent fluorescence spectroscopy and atomically detailed simulations. Both experiment and simulation reproduced the qualitative trend and suggested that the fastest permeation occurred for positively charged tryptophan. Molecular dynamics simulations revealed that the translocation mechanism was assisted by a local defect and the permeation process was insignificantly influenced by the long-range electrostatic interactions, such as the membrane dipole potential.