Biophysical Characterisation of Globins and Multi-Heme Cytochromes Using Electron Paramagnetic Resonance and Optical Spectroscopy
by Desmet, Filip, Ph.D., UNIVERSITEIT ANTWERPEN (BELGIUM), 2011, 231 pages; 3485078

Abstract:

Heme proteins of different families were investigated in this work, using a combination of pulsed and continuous-wave electron paramagnetic resonance (EPR) spectroscopy, optical absorption spectroscopy, resonance Raman spectroscopy and laser flash photolysis.

The first class of proteins that were investigated, were the globins. The globin-domain of the globin-coupled sensor of the bacterium Geobacter sulfurreducens was studied in detail using different pulsed EPR techniques (HYSCORE and Mims ENDOR). The results of this pulsed EPR study are compared with the results of the optical investigation and the crystal structure of the protein. The second globin, which was studied, is the Protoglobin of Methanosarcina acetivorans, various mutants of this protein were studied using laser flash photolysis and Raman spectroscopy to unravel the link between this protein's unusual structure and its ligand-binding kinetics. In addition to this, the CN -bound form of this protein was investigated using EPR and the influence of the strong deformation of the heme on the unusual low gz values is discussed. Finally, the neuroglobins of three species of fishes, Danio rerio, Dissostichus mawsoni and Chaenocephalus aceratus are studied. The influence of the presence or absence of two cysteine residues in the C-D and D-region of the protein on the EPR spectrum, and the possible formation of a disulfide bond is studied.

The second group of proteins that were studied in this thesis belong to the family of the cytochromes. First the Mouse tumor suppressor cytochrome b561 was studied, the results of a Raman and EPR investigation are compared to the Human orthologue of the protein. Secondly, the tonoplast cytochrome b561 of Arabidopsis was investigated in its natural form and in two double-mutant forms, in which the heme at the extravesicular side was removed. The results of this investigation are then compared with two models in literature that predict the localisation of the hemes in this family of cytochromes. Finally the preliminary results of a detailed HYSCORE study of the four hemes in the cytochrome c3 of Desulfovibrio desulfuricans ATCC 27774 are presented.

 
AdviserSabine Van Doorslaer
SchoolUNIVERSITEIT ANTWERPEN (BELGIUM)
SourceDAI/B 73-02, p. , Dec 2011
Source TypeDissertation
SubjectsBiochemistry; Molecular physics; Biophysics
Publication Number3485078
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