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Title:Lateral organization and dynamics of phospholipids in membrane
Author(s):Wang, Suke
Doctoral Committee Chair(s):Glaser, Michael
Department / Program:Biochemistry
Degree Granting Institution:University of Illinois at Urbana-Champaign
Subject(s):Chemistry, Biochemistry
Abstract:Studies of the lateral organization and dynamics of phospholipids were carried out using fluorescence techniques and enzymatic methods. Using resonance energy transfer, no specific interactions were observed between gramicidin and dansyl-labeled phosphatidylcholine, phosphatidylethanolamine, or phosphatidic acid. However, another integral membrane protein, D-$\beta$-hydroxybutyrate dehydrogenase (BDH), reorganized the phospholipids in the bilayer into a nonrandom distribution. Although the enzyme has a specific requirement for phosphatidylcholine for activity, the extent of energy transfer decreased in the order phosphatidic acid, phosphatidylcholine, phosphatidylethanolamine.
A strong interaction between BDH and phosphatidic acid was also observed with enzymatic methods. Phosphatidylcholine fully activated BDH only in the presence, but not in the absence, of phosphatidic acid in the vesicle. This is consistent with a mechanism where phosphatidic acid enhances the binding of BDH to membranes.
A redistribution of both gramicidin and dansyl-labeled phospholipids was easily observed when a phase separation was induced by adding Ca$\sp{2+}$ to vesicles made up of phosphatidylcholine and phosphatidic acid. Energy transfer measurements from gramicidin to either dansyl-phosphatidylcholine or dansyl-phosphatidic acid showed gramicidin preferentially partitioned into the phosphatidylcholine-rich regions. In the same system, two distributed fluorescence lifetimes or rotational correlation times were required to fit the data acquired with phase fluorometry for the dansyl-phospholipids and dansyl-gramicidin. Some of the dynamic parameters varied in different lipid domains. BDH appeared to partition into phosphatidylcholine-rich regions as detected by the changes in the Michaelis-Menten constants.
The orientations of diphenyl-hexatriene (DPH) in vesicle bilayers were studied using phase fluorometry and global analysis. The distributions were wide in the fluid phases and they narrowed dramatically in the gel phases. By linking the time zero anisotropy (r$\sb{\rm o}$) of several sets of data taken at various temperatures in a single global analysis, between recovery of the rank order parameters, the diffusion constant and r$\sb{\rm o}$ were obtained. The results suggest that a single distributed population of DPH was present in the bilayers with their orientational distributions dependent upon the physical state of the membrane.
Issue Date:1989
Rights Information:Copyright 1989 Wang, Suke
Date Available in IDEALS:2011-05-07
Identifier in Online Catalog:AAI9011070
OCLC Identifier:(UMI)AAI9011070

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