STUDIES ON THE EXCRETED ALKYL SULFATE MEMBRANE OF OCHROMONAS DANICA.
Item
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Title
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STUDIES ON THE EXCRETED ALKYL SULFATE MEMBRANE OF OCHROMONAS DANICA.
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Identifier
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AAI8222943
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identifier
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8222943
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Creator
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EL-MARAGHY, EZZAT B.
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Contributor
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Thomas H. Haines
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Date
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1982
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Language
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English
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Publisher
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City University of New York.
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Subject
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Chemistry, Biochemistry
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Abstract
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The isolation and purification of the extracellular vesicles {lcub}ECV{rcub} of the phytoflagellate, Ochromonas danica, by two procedures is described. The procedures were simple, rapid and produced a pure membrane preparation. The isolated ECV was free of other cellular organelles as shown by electron microscopy. The membrane preparation consisted of uniformly sized vesicles (preparation I), with a trilamellar pattern. The preparation was analyzed for its lipid, protein and carbohydrate composition. The lipids of the ECV were nearly identical to those of the flagella membrane. The polar lipid did not contain phospholipids and were exclusively the chlorosulfolipids. These were about 70% of the total lipids, the remainder was non-polar lipids.;These sulfolipids have been previously characterized as 1,14-docoanediol-1,14-disulfate and 1,15-tetracosanediol-1,15-disulfate containing up to six chloro groups replacing hydrogen on the otherwise saturated hydrocarbon chain.;The non-polar lipids (total 22%) were dominated by a large amount of free fatty acid ((TURN)14%), and sterols ((TURN)7%).;The ECV were subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis and found to contain 4 major protein components. These had apparent M(,R) of 97,000, 85,000, 47,000 and 19,000. In addition one minor protein of apparent molecular weight of 110,000 dalton was found. All protein components were shown by periodic-acid-Schiff reagent to be glycoproteins.;The analyses showed that carbohydrate dominates the ECV ((TURN)33%). The major neutral sugar is mannose.;The charge distribution on the surface of the ECV was examined by the use of the alkylated pH indicators 4-heptadecyl-p-aminocoumarin and 4-undecyl-p-hydroxycoumarin. Using these indicators it was possible to measure the pH within 1.0 nm the of membrane surface. The results show that the membrane surface has a negative charge typical of model alkyl sulfate membranes, providing direct evidence that the alkyl disulfates behave as membrane polar lipids.;A study of the capacity of alkaline pH and sonication to dissolve this unique membrane indicates that the ECV alkyl disulfate membrane is stabilized by a high proton concentration.;Bovine serum albumin (BSA) was successfully used to remove some of the free fatty acids verifying their presence in the natural ECV membrane. The formation of multilamellar myelin-like membranes provoked by BSA treatment was demonstrated by electron microscopy.
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Type
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dissertation
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Source
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PQT Legacy CUNY.xlsx
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degree
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Ph.D.
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Program
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Biochemistry
