Use this URL to cite or link to this record in EThOS: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.795803
Title: Immunological and biosynthetic studies on the mammalian 2-oxoglutarate dehydrogenase complex
Author: Hunter, Anne
Awarding Body: University of Glasgow
Current Institution: University of Glasgow
Date of Award: 1985
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Abstract:
High, titre monospecific polyclonal antisera have been raised against purified mitochondrial 2-oxoglutarate dehydrogenase complex (OGDC) and its component enzymes, 2-oxoglutarate dehydrogenase (E1), lipoyl succinyltransferase (E2) and lipoamide dehydrogenase (E3). These specific antisera have been employed to monitor molecular events in the biosynthesis, import and maturation of this multimeric assembly. In antiserum raised against native, intact 2-oxoglutarate dehydrogenase complex, lipoamide dehydrogenase elicits a poor antibody response in comparison to the other polypeptides of the complex. It is thought this may reflect the conserved nature of lipoamide dehydrogenase which is involved in highly differing subunit interactions with the distinctive lipoyl acyltransferase (E2) 'core' enzymes of each of the individual 2-oxo acid dehydrogenase complexes. In cultured porcine kidney (PK-15) and bovine kidney (NBL-1) cells, incubated with [35S]methionine in the presence of uncouplers of oxidative phosphorylation, appearance of higher Mr forms of the individual enzymes can be detected by specific immune precipitation and fluorographic analysis. In the cases of 2-oxoglutarate dehydrogenase, E1, and lipoamide dehydrogenase, E3, the initial translation products have subunit Mr values 1000-3000 greater than the mature enzyme while the precursor of lipoyl succinyltransferase, E2, contains an additional sequence of Mr 6000-8000. Competition studies have revealed the immunological similarity of the precursor molecules to the native subunits. The precursor form of lipoyl succinyltransferase (pre-E2) is located in the cytoplasm of the cell prior to its conversion to the mature form and import into mitochondria. These precursor molecules are relatively stable, remaining in the cell for several hours if the cells are maintained in the presence of uncouplers. On removal of uncouplers, processing is rapidly initiated and is complete within 40 min. Interestingly antiserum to native 2-oxoglutarate dehydrogenase complex fails to recognise E2 precursor molecules which can be immunoprecipitated, however, by antibodies raised against the denatured E2 subunit. It is concluded that pre-E2 is conformationally dissimilar to native E2 which exists normally as a highly-ordered, multimolecular aggregate in the native complex.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.795803  DOI: Not available
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