Mossbauer and electron paramagnetic resonance studies of the cytochrome bf complex
Mossbauer and electron paramagnetic resonance studies of the cytochrome bf complex
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DOI：
10.1021/bi990080n
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发表时间：
1999-07-13
期刊：
影响因子：
2.9
通讯作者：
Haehnel, W
中科院分区：
文献类型：
--
作者：
Sch端nemann, V;Trautwein, AX;Haehnel, W
Fe-57富集的细胞色素bf复合物已被分离出菠菜的水培。它已被研究在不同的氧化还原状态，光学，EPR和穆斯堡尔谱。在190 K下，所有铁中心都处于氧化态的天然络合物的穆斯堡尔谱揭示了四种不同铁位点的存在：细胞色素B中的低自旋三价铁[异构体位移（δ）为0.20 mm/s，四极分裂（Δ E-Q）为1.77 mm/s，相对面积为40%]，细胞色素f的低自旋三价铁（Δ = 0.26 mm/s，Δ E-Q = 1.90 mm/s，相对面积为20%），和Rieske铁-硫蛋白（ISP）的两个高自旋三价铁位点与双半胱氨酸和双组氨酸连接的铁（δ（1）= 0.15mm/s，Δ E-Q1 = 0.70 mm/s，相对面积为20%，以及Δ（2）= 0.25 mm/s，Δ E-Q2 = 0.90 mm/s，相对面积为20%）。EPR和磁性穆斯堡尔测量在低温下证实了这些结果。对EPR数据和磁穆斯堡尔谱数据的晶体场分析得出了细胞色素B（3.60、1.35和1.1）和细胞色素f（3.51、1.69和0.9）的g张量（g（z）、g（y）和g（x））的估计值。抗坏血酸的加入不仅将细胞色素f的铁还原为亚铁低自旋态（在4.2 K下δ = 0.43 mm/s，Δ E-Q = 1.12 mm/s），而且将Rieske 2Fe-2S中心的双组氨酸配位铁还原为亚铁高自旋态（在4.2 K下δ（2）= 0.73 mm/s，Δ E-Q 2 = -2.95 mm/s）。在这个氧化还原步骤中，细胞色素B的穆斯堡尔参数没有改变，表明细胞色素f和Rieske蛋白的氧化还原变化没有改变细胞色素B中低自旋三价铁的第一配体球。用连二亚硫酸盐还原进一步将细胞色素B的两个血红素还原为亚铁低自旋态（在4.2 K下，Δ = 0.49 mm/s，Δ E-Q 1.08 mm/s）。在4.2K下磁性穆斯堡尔谱的自旋哈密顿分析产生了细胞色素bf复合物中还原的Rieske 2Fe-2S中心的超精细参数，其非常类似于来自嗜热栖热菌的Rieske中心的那些报道[Fee，J.A.，Findling，K. L.，吉田，T.，等人（1984）J.Biol.Chem.259，124-133]。
The Fe-57-enriched cytochrome bf complex has been isolated from hydrocultures of spinach. It has been studied at different redox states by optical, EPR, and Mossbauer spectroscopy. The Mossbauer spectrum of the native complex at 190 K with all iron centers in the oxidized state reveals the presence of four different iron sites: low-spin ferric iron in cytochrome b [with an isomer shift (delta) of 0.20 mm/s, a quadrupole splitting (Delta E-Q) of 1.77 mm/s, and a relative area of 40%], low-spin ferric iron of cytochrome f (delta = 0.26 mm/s, Delta E-Q = 1.90 mm/s, and a relative area of 20%), and two high-spin ferric iron sites of the Rieske iron-sulfur protein (ISP) with a bis-cysteine and a bis-histidine ligated iron (delta(1) = 0.15 mm/s, Delta E-Q1 = 0.70 mm/s, and a relative area of 20%, and delta(2) = 0.25 mm/s, Delta E-Q2 = 0.90 mm/s, and a relative area of 20%, respectively). EPR and magnetic Mossbauer measurements at low temperatures corroborate these results. A crystal-field analysis of the EPR data and of the magnetic Mossbauer data yields estimates for the g-tensors (g(z), g(y), and g(x)) of cytochrome b (3.60, 1.35, and 1.1) and of cytochrome f (3.51, 1.69, and 0.9). Addition of ascorbate reduces not only the iron of cytochrome f to the ferrous low-spin state (delta = 0.43 mm/s, Delta E-Q = 1.12 mm/s at 4.2 K) but also the bis-histidine coordinated iron of the Rieske 2Fe-2S center to the ferrous high-spin state (delta(2) = 0.73 mm/s, Delta E-Q2 = -2.95 mm/s at 4.2 K). At this redox step, the Mossbauer parameters of cytochrome b have not changed, indicating that the redox changes of cytochrome f and the Rieske protein did not change the first ligand sphere of the low-spin ferric iron in cytochrome b. Reduction with dithionite further reduces the two hemes of cytochrome b to the ferrous low-spin state (delta = 0.49 mm/s, Delta E-Q 1.08 mm/s at 4.2 K). The spin Hamiltonian analysis of the magnetic Mossbauer spectra at 4.2 K yields hyperfine parameters of the reduced Rieske 2Fe-2S center in the cytochrome bf complex which are very similar to those reported for the Rieske center from Thermus thermophilus [Fee, J. A., Findling, K. L., Yoshida, T., et al. (1984) J. Biol. Chem. 259, 124-133].