Determining the Conformation of Apolipoprotein E4 in Spherical High-Density Lipoprotein by Crosslinking and Fluorescence Spectroscopy
<p> Apolipoprotein E is a 299-amino acid protein with two structural domains: an NT (1-191) and C-terminal (201-299) domain that bear high affinity binding sites to the LDL receptor and lipid, respectively. In the plasma and brain, apoE associates with phospholipids and cholesterol to form dis...
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| Language: | EN |
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California State University, Long Beach
2017
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| Online Access: | http://pqdtopen.proquest.com/#viewpdf?dispub=10635137 |
| Summary: | <p> Apolipoprotein E is a 299-amino acid protein with two structural domains: an NT (1-191) and C-terminal (201-299) domain that bear high affinity binding sites to the LDL receptor and lipid, respectively. In the plasma and brain, apoE associates with phospholipids and cholesterol to form discoidal nascent HDL which are remodeled into spherical HDL by the action of lecithin cholesterol acyltransferase (LCAT). We examined the conformation of apoE4 in spherical-HDL generated by the action LCAT on discoidal reconstituted HDL (rHDL) bearing single Cys variants employing fluorescence spectroscopy and site-specific crosslinking. The site-specific crosslinking studies showed the absence of intermolecular dimers which would be formed if two spatially proximal molecules are crosslinked. We also observed a loss of excimer between pyrene molecules attached covalently to single Cys variants of apoE4. The absence of dimers and loss of excimer indicate that apoE4 molecules in spherical rHDL are oriented in an anti-parallel fashion. This study provides more insight into the conformation of apoE4 in spherical HDL.</p><p> |
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