1H NMR spectroscopy of Paracoccus denitrificans cytochrome c-550.
1H NMR spectroscopy of Paracoccus denitrificans cytochrome c-550.
复制标题
脱氮副球菌细胞色素 c-550 的 1H NMR 光谱。
DOI：
10.1021/bi00310a022
复制
发表时间：
1984
期刊：
影响因子：
2.9
通讯作者：
Taylor,PV
中科院分区：
文献类型：
--
作者：
Timkovich,R;Cork,MS;Taylor,PV
Materials and MethodsParacoccus denitrificans (ATCC 13543) was cultured and cytochrome c-550 was isolated as described by Scholes et al.(1971). A finalchromatography step on hydroxyapatite was included in the purification (Ambler et al., 1981). The purity ratio (absorbance at the Soret maximum toabsorbance at 280 nm for the oxidizedform) of the final material was 5.2. This is equivalent to the value that may be calculated from the spectrum of pure cytochrome given in Scholes et al.(1971). In some experiments, material with a purity ratio of 4 was employed. Spectra of this material did not differ in the critical spectral regions of interest fromfully purified cytochrome. Samples for NMR were dialyzed vs. 50 mM ammonium bi-carbonate, pH 7.8, lyophilized, and redissolved in deuterated buffer. This solvent was 99.8% deuterium oxide, 10 mM potassium phosphate, and 100 mM sodium chloride, adjusted to the appropriate pH. Values labeled pH* represent the direct reading of a glass combination electrode in deuterium oxide after the electrodehad been calibrated in protic reference solutions. Adjustmentsof pH for titration studies were made by the addition of aliquots of 2HC1 or Na02H. Potassium cyanide and sodium azide were added to protein samples from concentrated stock solutions in 2H2G adjusted to the same pH* as the protein. Cytochrome reduction was accomplished by the addition of a minimum amount of solid sodium dithionite.
