Direct Measurement of the Effect of Cholesterol and 7-Dehydrocholesterol on Membrane Dipole Electric Field in Single and Mixed Sterol Vesicles Using Vibrational Stark Effect Spectroscopy

Cholesterol is an important contributor to the properties, structure, and function of biological membranes. An immediate biosynthetic precursor to cholesterol, 7-dehydrocholesterol (7DHC), differs only in a single double bond, yet defects in the conversion of 7DHC to cholesterol result in metabolic disorders. The membrane dipole field, <i>F⃗</i>_d, is the greatest contributor to the total membrane electric field and arises from the alignment of interfacial lipids and water molecules. We have previously shown, using vibrational stark effect (VSE) spectroscopy, that the magnitude of membrane dipole field decreases with increasing cholesterol content and that sterol structure can differentially affect <i>F⃗</i>_d. Here, we employ VSE spectroscopy to directly measure <i>F⃗</i>_d in small unilamellar vesicles (SUVs) composed of DMPC and 0-40% cholesterol or 7DHC. Our results show that cholesterol and 7DHC influence <i>F⃗</i>_d in a similar way, reducing the magnitude of <i>F⃗</i>_d with increasing sterol content in a trimodal manner corresponding to phase transitions in DMPC/sterol bilayers. To probe <i>F⃗</i>_d in membranes that better model diseased states, VSE measurements were performed on SUVs with 40% total sterol, but mixed compositions of cholesterol and 7DHC. <i>F⃗</i>_d was bimodal in these systems, reflecting a 7DHC-like field at 10% cholesterol/30% 7DHC but a cholesterol-like content at 15% cholesterol (25% 7DHC) and above. We propose possible sources of these trimodal and bimodal effects and discuss the implications on our understanding of membrane electrostatics and cholesterol-related metabolic disorders.