The ubiquinone pool and ubiquinol-cytochrome c oxidoreductase (cytochrome bc1 or Complex III) are central to cellular energy conversion in a very broad range of biological systems. Complex III converts oxidation- reduction (redox) free-energy stored in the substrates ubiquinol and cytochrome c into a transmembrane electric potential and pH gradient essential for cellular maintenance and, because of its reversibility, mitochondrial regulation. However, the Qo site of primary energy conversion in Complex III is poorly defined in crystal structures and experimentally difficult to access. We use molecular biology to excise all cofactors of the high and low potential chains except those that allow us to access to the molecular mechanisms of Complex III. These cofactor `knockouts'in the photosynthetic bacterium Rhodobacter capsulatus, are studied in vivo or in isolated reaction center/Complex III assemblies. The knockouts permit selective, controlled light generation of key catalytic states of reversible Qo site oxidation-reduction and coupled energy conversion. These quasi- equilibrium catalytic states are constrained to the Qo site, generated in high yields and persist for seconds at room temperatures. We expect in certain cases that they can be stabilized indefinitely. It is now viable to investigate these newly isolated catalytic states with a range of conventional and advanced structural, spectroscopic and electrochemical methods. These include infrared (ATR-FTIR) and electron paramagnetic resonance (EPR) to describe redox structures and mechanistic chemistry of the Qo site cofactors, quinone/quinol and recently, possibly the semiquinone radical in the quasi-equilibrium states. We hope to crystallize stabilized quasi-equilibrium Qo site states for X-ray structures of mechanistically relevant structures not considered possible before. Moreover, the lifetime of the quasi-equilibrium states lends each of them to be individually tested for reactivity with molecular oxygen with a view to identify mechanism and role in superoxide radicals and health critical ROS generation. All approaches promise novel molecular level descriptions of functional involvement of critical amino acids during active quinone-quinol binding and exchange, quinone- quinol oxidation-reduction and coupled charge separation catalysis. We believe that the approaches will also provide insight into the defensive strategies adopted by the Qo site for suppression of short circuits as well into the regulation of ROS. Experiments aided by general application of electron tunneling expressions will explore the underlying engineering of the Complex III, and other respiratory complexes for normal operation in forward and reverse modes, as well as determining the thresholds of failure and when destructive side reactions, including ROS generation, become significant. All aspects of the work will help bridge the significant gap between respiratory mechanism/energetics and mitochondrial and hence cellular regulation.
The energy obtained by our food and breath is managed at the molecular level by a set of specific proteins that produce and control the high energy chemicals essential for life.
We aim to understand how this molecular machinery controls such reactive chemicals as oxygen radicals to allow, on the one hand, normal healthy development and growth, and to minimize, on the other hand, the cellular damage that leads to aging and age-related diseases.
| Swierczek, Monika; Cieluch, Ewelina; Sarewicz, Marcin et al. (2010) An electronic bus bar lies in the core of cytochrome bc1. Science 329:451-4 |
| Zheng, Zhong; Dutton, P Leslie; Gunner, M R (2010) The measured and calculated affinity of methyl- and methoxy-substituted benzoquinones for the Q(A) site of bacterial reaction centers. Proteins 78:2638-54 |
| Chobot, Sarah E; Zhang, Haibo; Moser, Christopher C et al. (2008) Breaking the Q-cycle: finding new ways to study Qo through thermodynamic manipulations. J Bioenerg Biomembr 40:501-7 |
| Zhang, Haibo; Chobot, Sarah E; Osyczka, Artur et al. (2008) Quinone and non-quinone redox couples in Complex III. J Bioenerg Biomembr 40:493-9 |
| Zhang, Haibo; Osyczka, Artur; Dutton, P Leslie et al. (2007) Exposing the complex III Qo semiquinone radical. Biochim Biophys Acta 1767:883-7 |
| Osyczka, Artur; Zhang, Haibo; Mathe, Christelle et al. (2006) Role of the PEWY glutamate in hydroquinone-quinone oxidation-reduction catalysis in the Qo Site of cytochrome bc1. Biochemistry 45:10492-503 |
| Zhang, Haibo; Osyczka, Artur; Moser, Christopher C et al. (2006) Resilience of Rhodobacter sphaeroides cytochrome bc1 to heme c1 ligation changes. Biochemistry 45:14247-55 |
| Moser, Christopher C; Farid, Tammer A; Chobot, Sarah E et al. (2006) Electron tunneling chains of mitochondria. Biochim Biophys Acta 1757:1096-109 |
| Iwaki, Masayo; Yakovlev, Gregory; Hirst, Judy et al. (2005) Direct observation of redox-linked histidine protonation changes in the iron-sulfur protein of the cytochrome bc1 complex by ATR-FTIR spectroscopy. Biochemistry 44:4230-7 |
| Osyczka, Artur; Moser, Christopher C; Dutton, P Leslie (2005) Fixing the Q cycle. Trends Biochem Sci 30:176-82 |
Showing the most recent 10 out of 56 publications
