Identification of histidine as an axial ligand to P700+.
Identification of histidine as an axial ligand to P700+.
复制标题
鉴定组氨酸作为 P700 的轴向配体。
DOI：
10.1021/bi961765f
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发表时间：
1996
期刊：
影响因子：
--
通讯作者：
Babcock,GT
中科院分区：
文献类型：
--
作者：
Mac,M;Tang,XS;Diner,BA;McCracken,J;Babcock,GT
光系统I（PSI）中的主要电子供体P700被认为是二聚体的Chlaspecies。阳离子自由基的电子结构和几何结构都不清楚。磁共振研究表明，P700+中的未成对电子在Chladimer上不对称地离域;然而，中心Mg 2+的轴向配体尚不清楚。最近开发的一个组氨酸耐受突变体的集胞藻PCC 6803，使我们能够使用同位素标记和电子核双共振（ENDOR）光谱的组合，以显示第一个明确的光谱证据的组氨酸配体P700+。在15 N标记的P700+和[15 N]组氨酸特异性标记的P700+的ENDOR谱中观察到了关于15 N Larmor频率对称分裂的峰，对应于0.64 MHz的各向同性超精细耦合。这些峰在“反向”标记的样品中消失，其中所有氮都是15 N，除了组氨酸的氮，组氨酸含有天然丰度的14 N。偶极贡献的超精细耦合，通过使用电子自旋回波包络调制光谱（ESEEM）确定。ESEEM数据的数值模拟表明，耦合主要是各向同性的，组氨酸是直接协调的中心Mg 2+的P700+。两者合计，这些数据是支持的模型P700+中的激发态分子轨道作出了重大贡献的自由基的电子结构。此外，在这项工作中开发的方法可以扩展到检查在各种生物相关的卟啉/二氢卟酚系统的轴向配体的磁性。
The primary electron donor in photosystem I (PSI), P700, is thought to be a dimeric Chlaspecies. Neither the electronic nor geometric structure of the cation radical is clearly understood. Magnetic resonance studies have indicated that the unpaired electron in P700+is delocalized asymmetrically over the Chladimer; however, the axial ligand to the central Mg2+is not known. The recent development of a histidine tolerant mutant ofSynechocystisPCC 6803 has allowed us to use a combination of isotopic labeling and electron nuclear double resonance (ENDOR) spectroscopy to show the first definitive spectroscopic evidence of a histidine ligand to P700+. Peaks split symmetrically about the15N Larmor frequency corresponding to an isotropic hyperfine coupling of 0.64 MHz were observed in the ENDOR spectra from P700+globally labeled with15N and specifically labeled with [15N]histidine. These peaks disappeared in “reverse” labeled samples in which all nitrogens are15N except those of histidine, which contains natural abundance14N. The dipolar contribution to the hyperfine coupling was determined by using electron spin echo envelope modulation spectroscopy (ESEEM). Numerical simulations of the ESEEM data suggest that the coupling is primarily isotropic and that the histidine is directly coordinated to the central Mg2+of P700+. Taken together, these data are supportive of a model of P700+in which the excited state molecular orbital makes a significant contribution to the electronic structure of the radical. Moreover, the methodology developed in this work can be extended to examine the magnetic properties of axial ligands in a variety of biologically relevant porphyrin/chlorin systems.