Purification and characterization of a neuronal nicotinic acetylcholine receptor from Drosophila melanogaster.
Item
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Title
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Purification and characterization of a neuronal nicotinic acetylcholine receptor from Drosophila melanogaster.
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Identifier
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AAI9119692
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identifier
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9119692
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Creator
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Wu, Peipei.
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Contributor
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Adviser: Thomas Schmidt-Glenewinkel
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Date
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1991
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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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Biology, Neuroscience | Biology, General | Biology, Molecular
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Abstract
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The central nervous system of Drosophila contains an {dollar}\alpha{dollar}-Bungarotoxin binding activity with the properties expected of a nicotinic acetylcholine receptor. The receptor was purified 5800-fold by affinity chromatography. The receptor has a S{dollar}\sb{lcub}\rm 20,w{rcub}{dollar} of approximately 9.5 and a Stoke's radius of 7.4 nm. The frictional coefficient was calculated to be 1.7, indicating a highly asymmetrical protein structure. From sedimentation analysis in H{dollar}\sb2{dollar}O and D{dollar}\sb2{dollar}O, a molecular weight of 270,000 dalton was determined. SDS polyacrylamide gel electrophoresis followed by silver staining revealed two subunits with apparent molecular weights of 44,000 and 57,000 daltons. Polypeptides of the same size were identified by photoaffinity labeling of the crude membrane fraction in situ and of the purified receptor protein. The individual polypeptides were blotted onto a nitrocellulose membrane, digested with trypsin, separated by microbore reverse phase HPLC and subjected to internal amino acid sequence analysis. Screening of a cDNA library from Drosophila with oligonucleotide probes derived from the amino acid sequence allowed identification of several cDNA clones.
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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.
