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Title: The assimilation of carbon dioxide by Clostridiom kluyveri.
Authors: Andrew, Ian Godfrey.
Award date: 1966
Presented at: University of Leicester
Abstract: Clostridiom kluyveri, growing anaerobically in a medium containing ethanol, acetate and bicarbonate as sole carbon sources, rapidly incorporated radioactivity from 14C-bicarbonate into a wide variety of cell constituents. The first two compounds to become appreciably labelled were alanine and aspartate : after 6 seconds' incubation, alanine contained 84% of the total incorporated radioactivity, but this percentage subsequently decreased, while that in aspartate and in most other compounds increased. This suggested that alanine was derived through the carboxylation of a non-radioactive 2-carbon precursor, any intermediates in its formation being of only transitory existence, while other evidence suggested that a second carboxylation was involved in the formation of aspartate. The formation of alaninein cell-free extracts of C.kluyveri required the presence of acetyl-CoA, bicarbonate, hydrogen, and certain cofactors, and evidence was obtained for the participation of three enzymes; hydrogenase, catalysing the reduction of ferredoxin with hydrogen; pyruvate synthase, catalysing the reductive carboxylation of acetyl-CoA to pyruvate, and requiring reduced ferredoxin, thiamine pyrophosphate, and a divalent cation for its activity; and alanine transaminase. Evidence was also obtained that the formation of aspartate required two enzymes: a typical biotin- and ATP-dependent pyruvate carboxylase, catalysing the formation of oxaloacetate from pyruvate; and aspartate trans- aminase. These various enzymic activities were separated by standard fractionation procedures, but were mostly unstable, and little overall purification was achieved. The decarboxylation of oxaloacetate by C.kluyveri extracts was stimulated by ATP, suggesting the participation of PEP carb-oxykinase. The formation of phosphoenolpyruvate in crude extracts was obscured by the presence of pyruvate kinase, which catalysed its hydrolysis. The sources of energy and reducing power for biosynthetic reactions in C.kluyveri were discussed. A complete tricarboxylic acid cycle could not be demonstrated in cell-free extracts, although certain of the enzymes were present, and evidence was obtained for their participation in the synthesis of glutamate.
Type: Thesis
Level: Doctoral
Qualification: Ph.D.
Rights: Copyright © the author. All rights reserved.
Appears in Collections:Theses, Dept. of English
Leicester Theses

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