Cytochrome c' from the purple photosynthetic bacterium Allochromatium vinosum (CCP) displays a unique, reversible dimer-to-monomer transition upon binding of NO, CO, and CN-. This small, four helix bundle protein represents an attractive model for the study of other heme protein biosensors, provided a recombinant expression system is available. Here we report the development of an efficient expression system for CCP that makes use of a maltose binding protein fusion strategy to enhance periplasmic expression and allow easy purifn. by affinity chromatog. Coexpression of cytochrome c maturase genes and the use of a heme-rich Escherichia coli strain were found to be necessary to obtain reasonable yields of cytochrome c'. Characterization using CD, UV-vis spectroscopy, and size-exclusion chromatog. confirms the native-like properties of the recombinant protein, including its ligand-induced monomerization.
|Journal||Biochemical and Biophysical Research Communications|
|Publication status||Published - 2005|