| Scope and Method of Study. Determine the structure of Rhodobacter sphaeroides cytochrome bc1 complex using the engineered Super Reductase double mutant and studying the interaction between cytochrome c1 head domain and its electron acceptor, cytochrome c2. Studies were done using a variety of genetic and biochemical techniques.;Findings and Conclusions. The critical importance of cytochrome bc1 complex has made it a target for numerous antibiotics, fungicides, and anti-parasitic agents. The elucidation of the molecular mechanisms underlying these phenomena requires a combination of experimental approaches and in particular, structural investigations that can provide a molecular framework for further experiments. Because of the importance of the bacterial cyt bc1 complex in functional studies, a high resolution structure has been actively pursued for many years. In this study, we report an engineered mutant of Rb. sphaoeroides cyt bc1 complex, the SR mutant-SuperReductase that conspicuously shows higher enzymatic activity and significant increase in protein stability as compared to the wild type. The discovery of such an active and stable protein provided a breakthrough in the structure determination of Rb. sphaeroides cyt bc1 complex, providing a vital understanding of the structural-functional relationship of the complex. Additionally, cyt c1 head domain, the site of interaction between the cyt bc 1 complex and its electron acceptor cytochrome c 2, is isolated and purified to study without ambiguity the nature of interaction between the two proteins using differential scanning calorimetry. |
