Chemical, physical and spectroscopic studies of two clostridial-type 2(4-iron-4sulphur) ferredoxins.
Item
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Title
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Chemical, physical and spectroscopic studies of two clostridial-type 2(4-iron-4sulphur) ferredoxins.
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Identifier
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AAI9009736
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identifier
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9009736
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Creator
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Gluck, Martin Ronald.
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Contributor
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Adviser: William V. Sweeney
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Date
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1989
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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 clostridial-type 8Fe ferredoxins contain two 4Fe-4S centers, and are responsible for the oxidation-reduction properties of these proteins. Although it was originally thought for many clostridial-type 8Fe ferredoxins, particularly from Clostridium pasteurianum, the apparent midpoint reduction potential (E{dollar}\sb{lcub}\rm m{rcub}{dollar}) was pH dependent, evidence is presented in the first part of this thesis that demonstrates the absence of such a pH dependent midpoint reduction potential.;The central portion of this thesis describes some physical, chemical and spectroscopic studies of reductively methylated Clostridium pasteurianum 8Fe ferredoxin. The results show that the N-terminal amine in this protein is in an ion pair in both the oxidized and reduced forms of the protein, and that this ion pair is likely to be homologous to the one observed in Peptococcus aerogenes, another 8Fe ferredoxin for which an x-ray structure is known. Disruption of the normal ion pair by methylation appears to substantially decrease the intrinsic stability of the protein. Also presented is a spectroscopic analysis of the optical purity ratio, A{dollar}\sb{lcub}390{rcub}{dollar}/A{dollar}\sb{lcub}280{rcub}{dollar}, that characterizes the quantitative meaning of this widely used ratio. By the methods described it is possible to correlate the extent of protein denaturation with the change in the purity ratio as the protein undergoes degradation.;The latter part presents a thorough investigation of the products formed when 8Fe ferredoxin from either Clostridium pasteurianum or Clostridium acidi-urici is reacted with potassium ferricyanide. The results indicate that a variety of products are formed which contain varied amounts of peptide, iron and sulfide. The different iron and sulfide content reflects the presence of various iron-sulfur centers including the 3Fe and 4Fe types. However, a more homogeneous protein is formed if the native protein is allowed to degrade under aerobic conditions.;The final part examines the uptake of isotopically prepared alpha-{dollar}\sp2{dollar}H-L-cysteine into the 8Fe ferredoxin from Clostridium acidi-urici so that assignments can be made for these resonances in the {dollar}\sp1{dollar}H-NMR spectrum of this protein. NMR analysis revealed that three of the eight alpha-cysteinyl resonances are downfield shifted while the remaining five appear to be buried within the aliphatic envelope.
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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.
