Raman spectroscopy of supported lipid bilayers and membrane proteins
<p>Off-resonance unenhanced total internal reflection (<em>TIR</em>) Raman Spectroscopy was explored to investigate supported single lipid bilayers with incorporated membrane peptides/proteins at water/solid interface.</p> <p>A model membrane was formed on a planar sup...
| Main Authors: | , |
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| Format: | Thesis |
| Language: | English |
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2005
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| _version_ | 1797076460874235904 |
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| author | Lee, C Lee, Chongsoo |
| author2 | Bain, C |
| author_facet | Bain, C Lee, C Lee, Chongsoo |
| author_sort | Lee, C |
| collection | OXFORD |
| description | <p>Off-resonance unenhanced total internal reflection (<em>TIR</em>) Raman Spectroscopy was explored to investigate supported single lipid bilayers with incorporated membrane peptides/proteins at water/solid interface.</p> <p>A model membrane was formed on a planar supported lipid layer (<em>pslb</em>) by the fusion of the reconstituted small unilamellar vesicles (<em>SUV</em>s), and the intensity of bilayer was confirmed by a comparison of Raman spectral intensity in the C-H stretching modes with C<sub>16</sub>TAB. With prominent Raman sensitivity attained, we studied the 2-<em>D</em> phase transition of <em>DMPC</em> and <em>DPPC pslbs</em> and the temperature-dependent polarised spectra revealed a broad transition range of ca. 10 °C commencing at the calorimetric phase transition temperature.</p> <p>We applied polarised <em>TIR</em>-Raman Spectroscopy to <em>pslbs</em> formed by <em>DMPC SUV</em>s reconstituted with a model membrane-spanning peptide gramicidin D. A preferential channel structure formed by dissolution of trifluoroethanol could be probed by polarised Raman Spectroscopy qualitatively showing an antiparallel β-sheet conformation (different from "standard" one) and our Raman spectra by correlation with <em>NMR</em> and <em>CD</em> data confirmed single-stranded π<sup>6.3</sup> β-helical channel structure in the single bilayer. We also studied the membrane-penetrating peptide indolicidin in the presence of <em>DMPC pslb</em> over the chain melting temperature and a β-turn structure was dominantly observed concomitant with membrane perturbation.</p> <p>Dynamic adsorption of <em>DPPC</em> to form <em>pslb</em> from a micellar solution of <em>n</em>-dodecyl-β- <sub>D</sub>-maltoside could be examined with high sensitivity of every 1-<em>min</em> acquisition. Finally we used polarised <em>TIR</em>-Raman scattering to porcine pancreatic phospholipase A<sub>2</sub> hydrolytic activity on <em>DPPC pslbs</em> and revealed lipid-active conformation different from that of the enzyme alone.</p> |
| first_indexed | 2024-03-07T00:04:06Z |
| format | Thesis |
| id | oxford-uuid:76f4be6e-b7d3-46c5-a2a1-3dcc7a399410 |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-03-07T00:04:06Z |
| publishDate | 2005 |
| record_format | dspace |
| spelling | oxford-uuid:76f4be6e-b7d3-46c5-a2a1-3dcc7a3994102022-03-26T20:19:54ZRaman spectroscopy of supported lipid bilayers and membrane proteinsThesishttp://purl.org/coar/resource_type/c_db06uuid:76f4be6e-b7d3-46c5-a2a1-3dcc7a399410Raman spectroscopyMembrane proteinsBilayer lipid membranesEnglishPolonsky Theses Digitisation Project2005Lee, CLee, ChongsooBain, CBain, C<p>Off-resonance unenhanced total internal reflection (<em>TIR</em>) Raman Spectroscopy was explored to investigate supported single lipid bilayers with incorporated membrane peptides/proteins at water/solid interface.</p> <p>A model membrane was formed on a planar supported lipid layer (<em>pslb</em>) by the fusion of the reconstituted small unilamellar vesicles (<em>SUV</em>s), and the intensity of bilayer was confirmed by a comparison of Raman spectral intensity in the C-H stretching modes with C<sub>16</sub>TAB. With prominent Raman sensitivity attained, we studied the 2-<em>D</em> phase transition of <em>DMPC</em> and <em>DPPC pslbs</em> and the temperature-dependent polarised spectra revealed a broad transition range of ca. 10 °C commencing at the calorimetric phase transition temperature.</p> <p>We applied polarised <em>TIR</em>-Raman Spectroscopy to <em>pslbs</em> formed by <em>DMPC SUV</em>s reconstituted with a model membrane-spanning peptide gramicidin D. A preferential channel structure formed by dissolution of trifluoroethanol could be probed by polarised Raman Spectroscopy qualitatively showing an antiparallel β-sheet conformation (different from "standard" one) and our Raman spectra by correlation with <em>NMR</em> and <em>CD</em> data confirmed single-stranded π<sup>6.3</sup> β-helical channel structure in the single bilayer. We also studied the membrane-penetrating peptide indolicidin in the presence of <em>DMPC pslb</em> over the chain melting temperature and a β-turn structure was dominantly observed concomitant with membrane perturbation.</p> <p>Dynamic adsorption of <em>DPPC</em> to form <em>pslb</em> from a micellar solution of <em>n</em>-dodecyl-β- <sub>D</sub>-maltoside could be examined with high sensitivity of every 1-<em>min</em> acquisition. Finally we used polarised <em>TIR</em>-Raman scattering to porcine pancreatic phospholipase A<sub>2</sub> hydrolytic activity on <em>DPPC pslbs</em> and revealed lipid-active conformation different from that of the enzyme alone.</p> |
| spellingShingle | Raman spectroscopy Membrane proteins Bilayer lipid membranes Lee, C Lee, Chongsoo Raman spectroscopy of supported lipid bilayers and membrane proteins |
| title | Raman spectroscopy of supported lipid bilayers and membrane proteins |
| title_full | Raman spectroscopy of supported lipid bilayers and membrane proteins |
| title_fullStr | Raman spectroscopy of supported lipid bilayers and membrane proteins |
| title_full_unstemmed | Raman spectroscopy of supported lipid bilayers and membrane proteins |
| title_short | Raman spectroscopy of supported lipid bilayers and membrane proteins |
| title_sort | raman spectroscopy of supported lipid bilayers and membrane proteins |
| topic | Raman spectroscopy Membrane proteins Bilayer lipid membranes |
| work_keys_str_mv | AT leec ramanspectroscopyofsupportedlipidbilayersandmembraneproteins AT leechongsoo ramanspectroscopyofsupportedlipidbilayersandmembraneproteins |