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dc.contributor.advisorZhu, Xiaoyang, 1963-en
dc.contributor.advisorWebb, Lauren J.en
dc.creatorHu, Wenhui, M.A.en
dc.date.accessioned2012-02-16T19:49:28Zen
dc.date.available2012-02-16T19:49:28Zen
dc.date.issued2011-12en
dc.date.submittedDecember 2011en
dc.identifier.urihttp://hdl.handle.net/2152/ETD-UT-2011-12-4643en
dc.descriptiontexten
dc.description.abstractThe electrostatic field created by the inward pointing dipole moments of an oriented membrane leaflet has never been measured directly, but is thought to have an important influence on membrane function. Here we present the first direct measurement of the membrane dipole field in lipid bicelles using vibrational Stark effect spectroscopy which is based on the sensitivity of a nitrile oscillator’s vibrational frequency to its local electrostatic environment. The nitrile probe was introduced as the artificial amino acid p-cyanophenylalanine (CN-Phe) in four different locations of a α-helical peptide composed of alternating alanine and leucine residues. This peptide was intercalated into bicelles composed of mixtures of the long chain lipids 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), or 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), and the short chain lipid 1,2-dihexanoyl-sn-glycero-3-phosphocholine (DHPC) formed in two different sizes, 5 nm and 15 nm in radius. Formation of the bicelle above the phase transition temperature of the lipid mixture was confirmed by ³¹P NMR, and the structure of the [alpha]-helix within the bicelle was confirmed by circular dichroic spectroscopy. The absorption energy of the nitrile probe at 4 positions along the helical axis was measured by Fourier transform infrared spectroscopy, from which we estimate the magnitude of the membrane dipole electrostatic field to be -6 MV/cm. Then we successfully manipulated the dipole field in q = 0.5 DMPC/DHPC bicelles by incorporating the small molecule phloretin into the membrane and measured the corresponding ratiometric fluorescence signal of the co-intercalated voltage gated dye di-8-ANEPPS. We measured 0.7 ± 0.2 cm⁻¹ blue shift in absorption energy of the nitrile probe due to the decrease in dipole field caused by phloretin, corresponding to a dipole field of -4.2 MV/cm. This change was essentially identical to what has been estimated through ratiometric fluorescence methods, indicating that VSE spectroscopy will be useful tool for measurement of the biological effects of electrostatic fields in lipid membranes.en
dc.format.mimetypeapplication/pdfen
dc.language.isoengen
dc.subjectVibrational Stark effecten
dc.subjectMembrane dipole potentialen
dc.subjectMembrane electrostatic fielden
dc.subjectBicelleen
dc.subjectDMPCen
dc.subjectDHPCen
dc.subjectp-cyanophenylalanineen
dc.titleProgress towards directly measuring the membrane dipole field in lipid bicelles using vibrational Stark effect spectroscopyen
dc.date.updated2012-02-16T19:49:36Zen
dc.identifier.slug2152/ETD-UT-2011-12-4643en
dc.description.departmentChemistryen
dc.type.genrethesisen
thesis.degree.departmentChemistryen
thesis.degree.disciplineChemistryen
thesis.degree.grantorUniversity of Texas at Austinen
thesis.degree.levelMastersen
thesis.degree.nameMaster of Artsen


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