Cytochrome bc1 (Complex III) is an important enzyme that takes part in the respiratory electron transport chain in vertebrates, yeast, and many bacteria. The complex exists as a dimer, in which each monomer contains three catalytic subunits: cytochrome c1, cytochrome b and the Rieske iron-sulfur protein or ISP. Within the inner mitochondrial membranes of eukaryotes, Complex III catalyzes the transfer of two electrons from ubiquinol (UQH2) to cytochrome c, a water-soluble protein, through a process called the modified Q-cycle mechanism. Under very specific conditions, such as mutations within cytochrome b, disruption of the normal mechanism leads to bypass reactions, including the formation of superoxide and reactive oxygen species. We have sought to restore the activity of a mutant of cytochrome b (T61V) by modifying UQH2, the natural substrate for this enzyme. The structure of the oxidized form, UQ consists of a p-quinone head group and a hydrophobic all-trans polyprenyl unit (tail) that can vary in length, depending on the species in which it is found. The present work highlights modifications to the substituent groups attached to the quinone head and to the all-trans polyprenyl tail. Since the midpoint potentials of these molecules are pH dependent, cyclic voltammetry and spectroelectrochemistry studies in buffered aqueous solutions have been carried out on these molecules (analogs of UQ10). Modifications of the substituent groups attached to the quinone head gave the molecules a different ability to either donate or receive electrons, while modifications to the length of the tail either increased or decreased the solubility of these molecules inside the phospholipid membrane. We examined the normal activity and the production of superoxide in wild-type and (T61V) of bacterial Rhodobacter sphaeroides in the presence of these analogs. We confirmed that to prevent damaging side reactions, normal operation of the Q-cycle requires a fairly narrow window of reduction potentials with respect to the ubiquinol substrate.
| Identifer | oai:union.ndltd.org:arizona.edu/oai:arizona.openrepository.com:10150/195424 |
| Date | January 2010 |
| Creators | Cedeno, Diana |
| Contributors | Walker, F. Ann, Walker, F. Ann, Mash, Eugene A., Walker, F. Ann, Glass, Richard S., Wysocki, Vicki H., Olenyuk, Bogdan Z. |
| Publisher | The University of Arizona. |
| Source Sets | University of Arizona |
| Language | English |
| Detected Language | English |
| Type | text, Electronic Dissertation |
| Rights | Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author. |
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