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Plasmid DNA Preparation> | … c y , a pUC57 plasmid containing the cycY gene with a C-terminal 10xHis-tag separated from the coding sequence by a TEV protease cleavage site was purchased from GenScript (Netherlands). This plasmid was transformed into E. coli JM109 harboring the pEC86 plasmid … | Get A Quote |
Respiration is carried out by a series of membrane-bound complexes in the inner mitochondrial membrane or in the cytoplasmic membrane of bacteria. Increasing evidence shows that these complexes organize into larger supercomplexes. In this work, we identified a supercomplex composed of cytochrome (cyt.) bc and aa-type cyt. c oxidase in Rhodobacter sphaeroides. We purified the supercomplex using a His-tag on either of these complexes. The results from activity assays, native and denaturing PAGE, size exclusion chromatography, electron microscopy, optical absorption spectroscopy and kinetic studies on the purified samples support the formation and coupled quinol oxidation:O reduction activity of the cyt. bc-aa sup... More