Download MendeleyThe effects of an engineered cation site on the structure, activity, and EPR properties of cytochrome c peroxidase.
Bonagura, C.A., Sundaramoorthy, M., Bhaskar, B., Poulos, T.L.(1999) Biochemistry 38: 5538-5545
- PubMed: 10220341
- DOI: https://doi.org/10.1021/bi982996k
- Primary Citation of Related Structures:
1JDR - PubMed Abstract:
Earlier work [Bonagura et al. (1996) Biochemistry 35, 6107] showed that the K+ site found in the proximal pocket of ascorbate peroxidase (APX) could be engineered into cytochrome c peroxidase (CCP). Binding of K+ at the engineered site results in a loss in activity and destabilization of the CCP compound I Trp191 cationic radical owing to long-range electrostatic effects. The engineered CCP mutant crystal structure has been refined to 1.5 A using data obtained at cryogenic temperatures which provides a more detailed basis for comparison with the naturally occurring K+ site in APX. The characteristic EPR signal associated with the Trp191 radical becomes progressively weaker as K+ is added, which correlates well with the loss in enzyme activity as [K+] is increased. These results coupled with stopped-flow studies support our earlier conclusions that the loss in activity and EPR signal is due to destabilization of the Trp191 cationic radical.
Organizational Affiliation:
Department of Molecular Biology & Biochemistry, University of California, Irvine 92697-3900, USA.
