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Higuchi M, Hirano Y, Kimura Y, Oh-oka H, Miki K, Wang ZY: Overexpression, characterization, and crystallization of the functional domain of cytochrome c (z) from Chlorobium tepidum. Photosynth Res. 2009 Oct;102(1):77-84.
Cytochrome c (z) is found in green sulfur photosynthetic bacteria, and is considered to be the only electron donor to the special pair P840 of the reaction center. It consists of an N-terminal transmembrane domain and a C-terminal soluble domain that binds a single heme group. Large scale expression of the C-terminal functional domain of the cytochrome c (z) (C-cyt c (z)) from the thermophilic bacterium Chlorobium tepidum has been achieved using the Escherichia coli expression system. The C-cyt c (z) expressed has been highly purified, and is stable at room temperature over 10 days of incubation for both reduced and oxidized forms. Spectroscopic measurements indicate that the heme iron in C-cyt c (z) is in a low-spin state and this does not change with the redox state. (1) H-NMR spectra of the oxidized C-cyt c (z) exhibited unusually large paramagnetic chemical shifts for the heme methyl protons in comparison with those of other Class I ferric cytochromes c. Differences in the (1) H-NMR linewidth were observed for some resonances, indicating different dynamic environments for these protons. Crystals of the oxidized C-cyt c (z) were obtained using ammonium sulfate as a precipitant. The crystals diffracted X-rays to a maximum resolution of 1.2 A, and the diffraction data were collected to 1.3 A resolution. |
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