Heme proteins, also termed cytochromes, are a widespread class of metalloproteins containing an Fe-protoporphyrin IX cofactor. They perform numerous functions in nature such as oxygen-transport by hemoglobin, monooxygenation reactions catalyzed by Cytochrome P-450, and electron transfer reactions during photosynthesis. The differences between proteincofactor binding characteristics and the cofactor environment greatly influence the extensive range of functions.
In this dissertation, proteins from the Mtr pathway of Shewanella oneidensis are characterized. These c-type cytochromes contain multiple heme cofactors per protein molecule that covalently attach to the protein amino acid sequence and are involved in electron transfer to extracellular metal oxides during anaerobic conditions. Successful recombinant expression of pathway components MtrC
and MtrA is achieved in Escherichia coli. Heme-dependent gel staining and UV/Vis spectroscopy show characteristic c-type cytochrome characteristics.
Mass spectrometry confirms that the correct extensive post-translational modifications were performed and the ten heme groups were incorporated per protein of MtrC and MtrA and the correct lipid-anchor was attached to extracellular MtrC. Raman spectroscopy measurements of MtrA provide
intriguing structural information and highlight the strong influence of the heme cofactors within the protein structure.
Next, an Arabidopsis thaliana protein is analyzed. It was previously identified via a motif search of the plant genome, based on conserved residues in the H4 NOX pocket. Here, the incorporation of a heme b cofactor is confirmed. UV/Vis spectroscopy under anaerobic conditions demonstrates reversible binding of nitric oxide to the heme iron and depicts the previously published characteristic
absorption maxima for other H-NOX proteins.
| Identifer | oai:union.ndltd.org:kaust.edu.sa/oai:repository.kaust.edu.sa:10754/627223 |
| Date | 01 1900 |
| Creators | Vogler, Malvina M. |
| Contributors | Rueping, Magnus, Biological and Environmental Sciences and Engineering (BESE) Division, Arold, Stefan T., Schwingenschlögl, Udo, Stingl, Ulrich |
| Source Sets | King Abdullah University of Science and Technology |
| Language | English |
| Detected Language | English |
| Type | Dissertation |
| Rights | 2019-03-05, At the time of archiving, the student author of this dissertation opted to temporarily restrict access to it. The full text of this dissertation became available to the public after the expiration of the embargo on 2019-03-05. |
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