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Struktur und Funktion humaner Acyl-CoA-Dehydrogenasen. Untersuchungen zum 'Sudden Infant Death Syndrom'

Struktur und Funktion humaner Acyl-CoA-Dehydrogenasen. Untersuchungen zum 'Sudden Infant Death Syndrom'

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KRÄUTLE, Franz-Georg, 2001. Struktur und Funktion humaner Acyl-CoA-Dehydrogenasen. Untersuchungen zum 'Sudden Infant Death Syndrom' [Dissertation]. Konstanz: University of Konstanz

@phdthesis{Krautle2001Struk-6953, title={Struktur und Funktion humaner Acyl-CoA-Dehydrogenasen. Untersuchungen zum 'Sudden Infant Death Syndrom'}, year={2001}, author={Kräutle, Franz-Georg}, address={Konstanz}, school={Universität Konstanz} }

deu The aim of this work was to establish an isolation method for recombinantly expressed human Acyl-CoA dehydrogenases form E. coli. The main interest was supposed to be the mutant K304E of medium-chain specific Acyl-CoA dehydrogenase. This enzyme may play an important role in the Sudden Infant Death.<br />An amino acid exchange at position 304 of the native enzyme, leads to a charge reversal an probably to an instability at this very position. Previous attempts to isolate this enzyme failed due to minute amounts or aggregation and degradation phenomena. A new method to detect and quantify recombinant Acyl-CoA dehydrogenase by the use of monospecific antiobodies was set up to unambiguosly measure inactive enzyme in crude extracts.<br />A chromatographic separation scheme based on maximum three different steps was developed to purify different forms of human Acyl-CoA Dehydrogenase. The mutant K304E of medim-chain specific acyl-CoA dehydrogenase could for the first time be separated in a highly pure form. Also, a transient binding of molecular chaperone GroEL could be shown for the mutant enzyme.<br />The biological activity of mutant K304E was against all expectations comparable to the native enzyme although the substrate specificity differed markedly towards long-chain fatty acid substrates. This could also be demonstrated by different extents of bleaching upon binding to substrates of various chain-lengths. The natural electron acceptor ETF also showed different binding behaviour to the mutant enzyme. All together, this lead to the assumption of slightly different shaped conformation of the mutant enzyme.<br />In analogy of the medium-chain specific enzyme, the recombinant long-chain specific enzyme could also be isolated using the same separation scheme. Long-chain specific Acyl-CoA dehydrogenase could crystalized and ist structure could be resolved. Struktur und Funktion humaner Acyl-CoA-Dehydrogenasen. Untersuchungen zum 'Sudden Infant Death Syndrom' application/pdf Structure and funktion of human Acyl-CoA dehydrogenases. Iinvestigations on the Sudden Infant Death Syndrome Kräutle, Franz-Georg Kräutle, Franz-Georg deposit-license 2011-03-24T17:30:23Z 2001 2011-03-24T17:30:23Z

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